US20080044426A1 (Novel Atypical Pneumonia-Causing Virus)

US 20080044426A1
(19) United States

(12) Patent Application Publication (10) Pub. No.: US 2008/0044426A1
De Jong et al. (43) Pub. Date: Feb. 21, 2008
(54) NOVELATYPICAL PNEUMONIA-CAUSING (30) Foreign Application Priority Data
VIRUS
Nov. 18, 2003 (EP) .................................. O3O78613.1
(76) Inventors: Jan Cornelis De Jong, Eh Gouda Dec. 1. 2003 (EP) --- ------ --- ----------- --------- -- O3O78772.5
(NL); Theodorus Marinus Publication Classification

Bestebroer, Ve Krimpen Aanden
Ijssel (NL); James Henry (51) Int. Cl.
Matthew Simon, Amsterdam A 6LX 39/25 (2006.01)
(NL); Ronaldus Adrianus Maria A 6LX 39/395 (2006.01)
Fouchier, Rotterdam (NL); A6IR 48/00 (2006.01)
Albertus Dominicus Marcellinus C07K 6/10 (2006.01)
Osterhaus, Antwerpen-Zuid (BE) CI2N I/2 (2006.01)
CI2N I5/40 (2006.01)
Correspondence Address: EN E. 308:
HOFFMANN & BARON, LLP CI2N 7/01 (2006.01)
6900 UERCHO TURNPIKE CI2N 9/12 (2006.01)
SYOSSET, NY 11791 CI2O 1/70 (2006.01)
(52) U.S. Cl. .................. 424/159.1; 424/221.1; 435/194;
(21) Appl. No.: 10/579,614 435/235.1: 435/252.3:435/348; 43.5/5: 514/2:
514/44; 530/350; 530/389.4:536/23.72
(22) PCT Filed: Nov. 18, 2004 (57) ABSTRACT
(86). PCT No.: PCT/NLO4/OO805 The invention relates to the field of Virology. The invention
provides a new isolated essentially mammalian positive
S 371 (c)(1), sense single stranded RNA virus (EMCR-CoV) within the
(2), (4) Date: Jul. 19, 2007 group of coronaviuses and components thereof.
Patent Application Publication Feb. 21, 2008 Sheet 1 of 87 US 2008/0044426A1
Fig. 1
AGATAGAGAATTTTCTTATTTAGACTTTGTGTCTACTCCTCTCAACTAAACGAAATTTTTCTAGTGCTGTCATTTGTTATGGCAGTCCTAGT
TCTATCTCTTAAAAGAATAAATCTGAAACACAGATGAGGAGAGTTGATTTGCTTTAAAAAGATCACGACAGTAAACAAT ACCGTCAGGATCA
Hmm.5'UTR mmm-H
GTAATTGAAATTTCCT CAATTTGTAAAcTGGTTAGGCAAGTGTTGTATTTCTSTGTAAGC AcTGGGTTCGTCC ACTAGTGCACAC
------ee-------------------------as-a------ee-us--------------------------------------- 84
CATTAACTTAAAGCAGTTCAAACATTTGACCAATCCGTTCACAACATAAAAGACACAAATCGTGACCACCAAGACAGGTGATCACGTGTG
mm. 5'UTR mummim

ATTGATACTTAAGTGGTGTTCTGTCACTGCTTATTGTGGAAGCAACGTTCTGTCGTTGTGGAAACCAATAACTGCTAACCATGTTTACAA
is re-as-Hee-s-s----a---------------------------------- 276
TAACTATGAATTCACCACAAGACAGTGACGAATAACACCTTCGTTGCAAGACAGCA ACACCTTTGGTTATTGACGATTGGTACAAAATGTA
m Replicase 1a
CAAGTGACACTTGCTGTTGCAAGTGATTCGGAAATTTCAGGTTTTGGTTTTGCCATTCCTTCTGTAGCCGTTCGCGCTTATAGCGAAGCCGC
------------------------------esse----------------------------------ee-ee------------------------- 36s
GTTCACTGTGAACGACAACGTTCACTAAGCCTTTAAAGTCCAAAACCAAAACGGTAAGGAAGACATCGGCAAGCGCGAATATCGCTTCGGCG
O W T L A v A S D S E i s G F G F A 1 P S v. A v R A Y S E A A
-Replicase 1a-H-
TGCACAAGGTTTTCAGGCAGCCGCTTTGTTGCTTTTGGCTTACAGGATTGTGTAACCGGTATTAATGATGACGATTATGTCATTGCATTGA
ACGGTTCCAAAAGTCCGTACGGCGAAACAACGAAAACCGAATGTCCTAACACATTGGCCATAATTACTACGCTAAACAGTAACGTAACT
A D G F O A C R F W A F G L O D C W T G N D D D Y V A L.
-Replicase ta
CGGTACTAATCAGCTTGTCCAAAATTTACTTTTCTGATAGACCTCTTAATGcGAGGTTGGCTCATTTTTCTAACAGCAATTAT
V - ---------------------------------------------------- 552
GACCATGATTAGTCGAAACACGGTTTTAAAATGAAAAAAGACTATCTGGAGAATTAAACGCTCCAACCGAGTAAAAAAGATTGTCGTTAATA
T G T N, O L C A K L. F S R P N R G W F s N S N Y
-Replicase 1 a
GTTCTTCAGGACTTTGATGTTGTTTTTGGCCATGGTGCAGGAAGTGTGGTTTTTGTGGATAAGTATATGTGTGGTTTTGATGGTAAACCTGT
------------------------------------------------------------------ 644
CAAGAAGTCCTGAAACTACAACAAAAACCGGTACCACGTCCTTCACACCAAAAACACCTATTCATATACACACCAAAACTACCATTTGGACA
V o F D W. V. F G H G A G S V V F W D K Y M C G F D G K P V
Replicase 1 a
GTTACCTAAAAACATGTGGGAATTTAGAGATTACTTTAATGATAATACTGATAGTATTGTTATTGGTGGTGTCACTTATCAATTAGCATGGG
-------------------------------------------------------------------------------------------- 736
CAAYGGATTTTTGTACACCCTTAAATCTCTAATGAAATTACTATTATGACTATCATAACAATAACCACCACAGTGAATAGTTAATCGTACCC
P K N M W E F R Y F N O N D S W G W Y O A W
Replicase 1 a
ATGT TATACGTAAAGACCTTTCT TATGAACAGCAAAATGTTTTAGCTATTGAGAGCATTCAT TATCTTGGCACTACAGGTCATACT TTGAAG
TACAAATGCATTCTGGAAAGAATACTGTCGTTTTACAAAATCGATAACTCTCGAAGTAATAGAACCGTAGCCATAAAACTTC
D v I R K D L S Y E O O N V Replicase
L. A 1a- E S H Y-
L G T T G H T L K
ACACAAGCTATTTGTGCTGCATTTGATTTTTCTTTAGATGTATTTAAAATTGGTGATGTTAAATTTAAACGACTTGGTGA TATGTTCTTAC
------------------------------------------------------------------------- 1748
TGTGTTCGATAAACACGACGTAAACTAAAAAGAAATCTACATAAATTTTAACCACTACAATTTAAATTTGCTGAACCACTAAT ACAAGAATG
T O A C A A F D F S L D v F K - G D V K F K R L G D Y v L. T
Replicase 1a- - -
TGAAAATGCTCTTGTTCGTTTGACTACTGAAGTTGTTCGTGGTGTTCGTGATGCTCGCATAAAGAAAGCCATGTTTACTAAAGTAGTTGTAG
---------------------------------ee-ee--------------------------------------- 1840
ACTTTTACGAGAACAAGCAAACTGATGACTTCAACAAGCACCACAAGCACTACGAGCGTATTTCTTTCGGTACAAATGATTTCATCAACATC
E N A W R L T T E W W R G W R D A R K. K. A. M. F. T. K. W. W. W
Replicase 1a
GTCCTACAACTGAAGT TAAGTTTTCTGTTATTGAACTTGCCACTGTAATTTGCGTCTTGTTGATTGTGCACCTGTAGTTTGCCCTAAAGGT
------------ee-ee-eels-----------------------elers---as-s-s-s-s-s-s-s-s-s-s-s-s-s-s-He--------------------------- 1932
CAGGATGTTGACTTCAATTCAAAAGACAATAACTTGAACGGTGACAATTAAACGCAGAACAACTAACACGTGGACAT CAAACGGGATTTCCA
G P T T E V K F is v I E L A T V N L R L v D C A P v v C P K G
Replicase 1a
AAAATTGTTGTTATTGCTGGACAAGCTTTTTTCTATAGTGGTGGTTTTTATCGTTTTATGGTTGATTCTACAACTGTATTAAATGACCCTGT
---------------------------------------------------------------- 2024
TTTTAACAACAATAACGACCGTTCGAAAAAAGATATCACCACCAAAAATAGCAAAATACCAACTA AGATGTGACATAATTACTGGGACA
K W W A G O A F F Y S G G F Y R F M W D S T T W L N D P W
Replicase 1 a
TTTTACTGGTGAGTTATTTTATACTATTAAGTTTAGTGGTTTTAAGCTTGATGGTTTTAACCATCAGTTTGTTAATGCTAGTTCTGCTACAG
-------------------------------------as--------------------------------------------------H 216
AAAATGACCACTCAAAAAAAGATAATTCAAATCACCAAAATTCGAACTACCAAAATGGTAGTCAAACAAT AGATCAAGACGATGT
E F Y T K F. S. G. F. K. L. D G F N H F W N A S S A T
Replicase 1a---
ATGCCATATTGCTGGAGCGTGTTATCGGATTTTAAAACTGCAGTTTTGTGACACAGTGGGTTGATGGTTGTAGTGTCATTGT
------------------------------------------ee------------ 2208
TACGGTAATAACGACAATCGACAACAATAGCCTAAAATTGACGTCAAAAACACAGTGTACACACCAACTACCAAATCACAGTAACAA
D A 1 A V E L L L S D F K T A. V. a F W Y T C v v D G C S v 1 v
-Replicase
AGACGTGATGCTACATTcGCCACACATGTGTGTTTTAAGGACTGTTATAGTATTTGGGAGCAATTCTGCATTGATAATTGTGGTGAGCCATG
------------------------------------------------------------------------------------------- 2300
TCTGACTACGATTAACGGTGTGTACACACAAAATCCTGACAAATCAAAACCCCGAAGACGTAACTATAACACCACTCGGTAC
R R D A T F A T H W C F K D C Y S W E O F C D N C G E P W
-Replicase 1a
GTTTAGATTATAATTACTTGCAGAGAATAACCCCAATTCATTTAAGCATCGGAGCTAAAGTTTTTGAGAGG
ass-s-s-s-s-s-s--------------------s-s-s-s-s-s-s-------------------------as-s-s-H- 2392
CAAAAACTGACTAATAACGATAAACGTCCATATTGGGAGTACACAAACAATTCGAGCCTCAGATTCAAAAGAACTTCCA
F D Y N A O S N N P A W A S E S K W L L E R
Replicase 1a- -
TTTTACCTAAGGTCCTGAAATACTGTGASTATTGAGATGGCCATTAGGAATCTTTTTTTGAAAAGTAATTTTGTTACAGATGG
AAAATGGATTCACAGGACTTTATGACAACTCATAACTACTACCGGT AAATACCTTAGAAAAACAACTTTTCAAATTAAAACAATGTCTAACC
P K C P E L L S D D, G H W N F W E K F N F W T D. W.
Replicase 1a
TTAAAAACTCTTAAGCT ACACTTACT CAATGGTTTTAGGTAATTGTGCCAAACGTAACGTTTTTGGTAAAATTCTIGATGT

---------------------------------------------------------------------------------- 2576
AATTTTAGAATTCGAATGTGAATGAAGATT ACCAGAAAATCATTAACACGGTTTGCAAAATCGCACAAAACATTTTAACGAACTACA
L K T - K L T L T S N G L L Replicase
G N 1Ca A K R F R R v L. v K L - D v
CTATAATGGTTTTCTTGAAACTGTCTGTAGTGTCGTACACACTGCTGGTGTTTGCATTAAATATTATGCTGTTAATGTTCCATATGTAGTTA
alles----------------------ener--------------------ee---------------gar-rre-pre-o-o-o-His-e-sa--all-a-ra-re 2668
GATATTAccAAAAGAACTTTGACAGACACACAGCAGTGTGACGACCACAAACGAATTTATAATACGACAATTACAAGGTATACATCAAT
Y N C F L E T W C S W W H A G V C K Y Y. A V N V P Y V V
Replicase 1 a
TAGTGGTTTGTAAGCGTGTAATCGT AGAGAAAGGGTGACGTGACTTTCCTTGTGTTAGTTGGTCACTTTTTCTATGAATTTTTA
----------------------------------------s-s-s-s-s-s-s-s-s-s-s-s-s-s-?-s-s-s-and-a--------------------------. 2760
AATCAccAAAACATTCAGCACATTAAGCATCTCTTTCCACACTGCACTGAAAAGGAACACAATCAACACAGTGAAAAAAGATACTTAAAAAT
S G F W S R v R R R C D W T F P C W S C W T F F Y E F
Replicase 1a
GACACGTTTTTGGTGFTAGTAAACTAATGCCATTGATGTTGAACATTAGAGCTTAAAGAAACTGTTTTTGTTGAACCTAAGGATGGTGG
------------------------------orres-s-s-s-s-Hers---------------------------------------- 2852
CT GTGCACAAAACCACAATCATTTGGATTACGGTAACTACAACTTGTAAATCTCGAATTTCTTTGACAAAAACAACTTGGATTCCTACCACC
T C F G V S K P N A V E H L E L K E T V F W E P K or G G
Replicase 1a
TCAATTTTTTTTTCTATAT TATCTTTGGTATGTTGTAGATGACATTTATTATCCAGCTTCATGTAATGGTGTATTGCCAGTTGCTTTTA
----------------------------------------------------------------------------- 2944
AGTAAAAAACAAAGACTACTAAT AGAAAccATAAACATTACTGTAAATAATAGGTCGAAGTACATTACCACATAACGGTCAACGAAAAT
O F F W S D D Y L w Y V V O D Y Y P A S C, N G V L. P. V. A F
Replicase 1a-------
CAAAATTGAGGGTAAAAATCTTTTCGATGATGTTATAGTTCATGATGTTGAACCTACCCAAAAGTCAAGCTCATATTTGAGTT
----------------------------------------------------------------------are------------------ 3036
GTTTTAACCGTCCACCATTAAGAAAAAGACACTACAATATCAAGTAcACAATTGAGTATTTATTGAGTATAAAccAAA
T K . A G G K s F s o o v i v H W E P T H K V K F E F
Replicase 1a
GAAGAGATGTTGTTACCAGTCTTGTAAGAAGAGTTTGGTAAGTCTATTATTATACAGGTGATGGGAAGGTTTACAGAAGTTTTAC
----------------------------------easter-----------ee-----------------------as-retre------------ 328
CTTCTACACAACAATGGTAGAAACATTTTCCAAAACCATTCAGATAATAAATAGTCCACTAACCCTTCCAAATsACTTCAAGAATG
E. D. D. W W T S C K K S F G K S Y T G W E G L H E W L
Replicase 1a- -
ATCTGCAATGAATGTCATTGGGCA ACATATTAAGTGCCACAATTTATATTTAGATGAAGAGGGTGGTTAGATGTTTCTA AACCAGTTA
-------------------------------------------------------------------------------as-e- 3220
TAACGTTACTTACAGTAACCCGTTGAAATCAACGGGTTAAAATATAAAACTACTTCTccCAccAATAcAAAAGATTGGTCAA
S A M N V G O H k l P O F Y Y E E G G Y V S K P V
-Replicase 1 a
TGATTCACAATGGCCTATAGTGATGATAGGATGGTTGTGFGTTGAAGCGAGC ACTSATTTCATAATTAGAATCTGTTAGAGAAGAG
----------------------------Hellas-e---ee------------------------------------------------- 332
ACT AAAGTGTT ACCGGATAAT CACTACTATCACT ACCAACACAACAACTTCGCTCGTGACTAAAAG TAGTTAACTTAGACAATCTCTTCTC
s o w P 1 s D D S D G C v V E aA
-Replicase S T D F H O L E S v R E E
GTTGAt ATAATTGAACAACCTTTTGGGGAAGTTGAACATGCGCTCT CAATTAGACAACCTTTTTCTTTTTCTTTTAGAGAIGAATTGGGTGT

3404
CAACTATATTAACTTGTTGGAAAACCCCTTCAACTTGTACGCGAGAGTT AATCTGTTGGAAAAAGAAAAAGAAAATCTCT ACTTAACCCACA
V D E O P F G E W E H A L S I R O P F S F is F R D E L G w
Replicase 1a- -- - -
TCGTGTTTTAGATCAATCTGATAATAATTGTTGGATTAGTACCACACTTATACAGTTGCAACTTACAAAGCTTTTGGATGATTCTATTGAGA
------------------------------------------------------------------------------ 34gs
AGCACAAAATCTAGTTAGACTATTATTAACAACCTAATCATGGTGTGAATATGT CAACGTTGAATGTTTCGAAAAccTACTAAGAT AACTCT
R v L. D. C S D N N C W E S T
-Replicase L aI O L O L T, K L L D D S E
GCAATTGTTAAAGTTGGTAAAGTGATTCAATTGTTCAAAAGTGTTAGAGTTGTCTCATTAATTAGGGTTCACTTGGGATAGTGGT
---------------------------------------------------a-a-a-ana-a----------------------- 3588
ACGTAACAAATTCAACCATTTCAACTAAGTAACAAGTTTCACAATACTCA ACAGAGTAAATTAACACCAAGTGAACCACTATCACCA
M. O. L. F K W G K W D S W 0 K Y E L S H S G S L G D S G
Replicase 1a
AAACTTCTTAGTGAACTTCTAAAGATAAATAACATGTTCTATAACTTTTGAGATGTCTTGGATGTGGTAAAAAGTTGAGAGCAAGT
------------------------------------------------------------------------------------- 3680
TTTGAAGAATCACTTGAAGAATTTCTATTTATATGTACAAGATATTGAAAACTCTACAGAACACTAACACCATTTTTCAAACTACTCGTTCA
K L L S E L L K K Y T C S T F E M S c C K K F D E O v
Replicase 1 a
TGGTTGTTTGTTTTGGATTATGCCTTACACAAAACTTTTTCAAAAAGGTGACTGTTCTATTTGTCATAAAATGCAGACTTATAAGCTTGTTA
---- e-ele--ee-s-s-s-s-s-s-s-s-s-s-----------Hire-to-r- 3772
ACCAACAAACAAAACCTAATACGGAATGTGTTTGAAAAAGTTTTCCACTCACAAATAAACAGTATTACGTCTGAATATTCGAACAAT
L. F W M P Y T K F K G E c c H K M C T Y K L v
Replicase 1a------,
GATAAAGTATTGTGTTTGACAGATCCAGCACCTATTGACATTGACTTTCCCGTAGACCTATATGTTCATCTGTATATTA
----------------------------------------------------------------------------- 3864
CATACTTTCCATGACCACACAAACATGTCCTAGGTCGTGGATAACTGTAACTACGAAAGGGACAATCTGGATATACAAGTAGACATATAAAT
S M K G T G W F W C o P A P D D A F P W R P c s s v Y
-Replicase ta
GGGTTAAGGGTTCTGGTCATTACAAACAAATTTAACAGTTTGACAAAGCTATTGAGGTTTGGTTCTTGACATAAAAAAGTAG
-------------------------------------------as-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s---------------------- 3956
CCACAATTCCCAAGACCAGTAATAGTTTGTTTAAAT ATGT CAAAACT GTTTCGATAACTACCAAAACCACAGAAACTGTAATTTTTATCATC
G W K. G. S G Y O N Y S F D K A D G F G W F o K N S S
Replicase 1 a
TGTTAATACTGTTTGTTTGTTGATGTTGATTTTCATAGTGTAGAAAT AGAAGCTGGTGAAGTTAAACTTTTGCTGTATATAAAAATGTTA
-----------------------her----------------------------ee-e-e-r-s-s-s-s-s-s-s-s-s-s-s-s--------------- OB
ACAATTAGACAAACAAAACAACTACAACTAAAAGTATACATCTATCTTCGACCACTTAATTTGGAAAAcGACATATATTACAAT
V N T W C F W W D F H S W E E A G E V K P F A V Y K N W
Replicase 1 a
AATTATTAGGGATATTTCACACCTTGTAAACTTGTCTTTGACTGTTGTCAATGCGCTAAGAAAATTCACAGAGGC
--------------sur-H-------------ee-as-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-H--------or-----------------one to
TTAAAATAAATCC ACTATAAAGTGGGAAATGAACAAAGAAAATGAAACAACAGTTACGACGATTACTTTTAGAGACGTACCTCC
K F Y G D S h W N C W S F D F W W N A A N E N M H G G
Replicase 1 a
Y GGTGTCGCACGTGCTATTGATATTTTGACTGAAGGTCAACTTCAGTCATTATCTAAAGATTACATTAGTAGTAATGGTCCACTTAAGGTTGG
-----------------------------------as-s-s-sa-i-lead-as-s-s-s-s-s-s-s-s--------- 4232
CCACAGCTGcACGATAACTATAAAACACTCCATTGAAGTCAGTAAT AGATTCTAATGTAATCATATTACAGGTGAATTCCAACC
G v A. R A D L T E G O L O S L S K D Y S S N G P L K V G
Replicase 1 a
AGCAGGTGTTATGTTGGAGTGTGAAAAATTCAATGTATTTAATGTTGTTGGTCCGCGAACTGGTAAACATGAGCATTCATT ACTTGTTGAAG
--------------------------------------------------------------------- 4324
TCTCCACAATACAACCT CACACTTTTTAAGTTACATAAATTACAACAACCAGGCGCTTGACCATTTGTACTCGTAAGTAATGAACAACTTC
A W M L E C E K F N W F N. W. W G P R T G K H E H S V E
Replicase a
CTTATAATTCTATTTTATTTGAAAATGGTATTCCACTTATGCCTCTTCTTAGTTGTGGTATTTTTGGTGTAAGGATTGAAAATTCTCTTAAA
s
GAATATTAAGATAAAATAAACTTTAccATAAGGTGAATAcGGAGAAGAATCAACACCAAAAAACCACATTCCTAACTTTTAAGAGAATT
A Y N S F E N G P L M P L L S C G F G W R E N S L K
-Repticase 1 a
GCTTTGTTTAGTTGTGACATTAATAAAccATTGCAAGTTTTTGTTTATTCTTCAAATGAAGAACAAGCTGTTC TTAAGTTTTTAGATGGTT
---------------------------------------------s-s-s-s-s-s-s-s-----------H 508
CGAAACAAATCAACACGTAAT ATTTGGTAACGTTCAAAAACAAATAAGAAGTTTACTTCTGTTCGACAAGAATTCAAAAACTACCAAA
A L. F s C D N K P L Q v F Replicase
v Y Sa S N E E O A v L K F L D G L
AGATTTAACACCAGTCATTGACGATGTTGATGTTGTTAAACCTTTTAGAGTTGAAGGTAATTTTTCATTC TTGATGGTTAATGCC
------------------------------------------------------eeee--------------------------- 600
Tct AAATTGTGGTCAGTAACTGCTACAACTACAACAATTTGGAAAATCTCAACTTCCATFAAAAAGTAAGAAACTAACACCAEAGTTACGGA
D L T P W D D v D V V K P F R W E G N F S F F O C G V N A
Replicase 1 a i
TGGATGGTGATATTT ACTTATTATTACTAACTCTATTTTAATGTTGGATAAACAAGGACAATTATTGGACACAAAACTTAATGGTATTTTG
----------------------------------------------eleas---------ele----------------ee-les---------------- 692
AccT ACCACTATAAATGAATAATAAATGATTGAGATAAAATTACAACCTATTTGTTCCTGTTAATAACCGTGTTTGAATTACCATAAAAC
D Y F T N S M K C G O L K L N G
Replicase 1a
CAACAGGCAGTTCTTGATTATCTTGCTACAGTTAAAACTGT ACCAGCTGGT AATTTGGTTAAACTTGTTGTTGAGAGTTGTACCATTTATAT
---------------------------------------------------Hea------------a-a-a-a-a-a-a-ra--a 784
GTTGTCCGTCAAGAACTAAT AGAAcGATGTCAATTTTGACATGGTCGACCATTAAACCAATTTGAACAACAACTCTCAACATGGTAAATATA
O O. A v L. D Y - A T V K T V P A G N L V K L V V E S C T
-e-r-Replicase 1 a
Y M
GTGTGTTGTACCATcGATAAATGATCTTTCTTTTGATAAAAATCTTGGTCGTTGTGTGCGTAAACTTAAT AGATTGAAAACTTGTGTTATTG
---------------------------------------------------------------------------- 4876
CACACAAATGGTAGCTATTT ACT AGAAAGAAAACTATTTTTAGAACCAGC AACACACGCATTTGAATTATCT AACTTTTGAACACAATAAC
C. v V P S N D L S F D K N L G R C W R K L N R K T C V
-ms-as-a-Replicase 1 a
CCAAGTTCCFGCTATTGATGTTTGAAAAAGCTCTTCAATTTGACTTTAACTGTAAATTTGTTGTAGAGAGTAATGTTATSGAGT
-----------------------------------------------------------em------------------- 4968
GGTACAAGGACGATAAC TACAAAACTTTTTCGAAGAAAGTTCAAACTGAAATTGACAATTTAAACAACATCTCTCATTACAATACCTACAA
A N V P A v L. K. k l S S T V K F W W E S N W M D W
Replicase 1a
AACGACTGTTTTAAGAATGATAATGTAGTTTTGAAAATTACTGAAGATGGTATTA ATGT TAAAGATGTfGTTGTTGAGTCTTCTAAGTCACT

-------------H------------------------------------------------------- 5060
TGCTGACAAAATTCTTACTATTACAT CAAAACTTTTAATGACTTCTACCAAATTACAATTTCTACAAcAACAACTCAGAAGATTCAGTGA
N D C F K N D N V V L. K. T E D G N V K D V V V E S S K S L.
Replicase 1a
TGGTAAACAATTGGGTGTTGTGAGTGATGGTGTTGACTCTTTTGAAGGTGTTTTACCTATTAATACTGATACTGTCTTATCTGTAGCTCCAG
H--------------------------------H-- 552
ACCATTTGTTAACCCACAACACTCACTACCACAACTGAGAAAACTTCCACAAAATGGATAATTATGACTATGACAGAATAGACATCGAGGTC
G K O L G V V S D G W D S F E G W L P N T D T W L S v A P
-Replicase 1a
AAGTTGACTGGGTTGCTTTTT ACGGTTTTGAAAAGGCAGCACTTTTTGCTTCTTTGGATGTAAAGCCATATGGTTACCCTAATGATTTTGTT
------------------------------------------------------------------------ 524.4
TTCAACTGACCCAACGAAAAATGCCAAAACTTTTCCGTCGTGAAAAACGAAGAAACC TACATTTCGGTATACCAATGGGATTACTAAAACAA
E v O. W W A F Y G F E K A A L. F A S L D V K P Y G Y P N D F V
Replicase 1 a
GGTGGTTTTAGAGTTCTTGGGACCACCGACAATAATTGTTGGGTTAATGCAACTTGTATAATTTTACAGTATCTTAAGCCTACTTTTAAATC
t-----------------------ee-a-H----------------------------- 5336
CCACCAAAATCTCAAGAACCCTGGTGGCTGTTATTAACAACCCAATTACGTTGAACATATTAAAATGTCATAGAATTCGGATGAAAATTTAG
G G F R W - G T T N N C W V N A T C L. O. Y L. K P T F K S
Replicase 1a
TAAGGGTTTAAATGTTCTTTGGAACAAATTTGTTACAGGTGATGTTGGACCTTTTGTTAGTTTTATTTATTTTATAACTATGTCTTCAAAGG
-a-m------------------------------------------------- 528
ATTCCCAAATTTACAAGAAACCTTGTTTAAACAATGTCCACTACAACCTGGAAAACAATCAAAATAAATAAAATATTGATACAGAAGTTTCC
K G L N V . W N K F W T G D W G P F W S F Y F | T M S S K
Replicase 1a
GTCAAAAGGGTGATGCGAAGAGCCATTATCTAAATTGTCAGAGTATTTGATTAGTGATTCTATTGTTACCTTGAACAATATTCAACTGT
H----------------------------------------------------------- 552O
CATTTCCCACTACGACTTCTCCGTAAT AGATTTAACAGTCTCATAAACTAATCACTAAGATAACAATGAGAACTTGTTATAATGAACA
G O K G D A E E A L S K S E Y S D S W T L E O Y S T C
-Replicase 1a
GACATTTGTAAAAGTACT GTAGTTGAAGTTAAAAGTGCTGTTGTCTGTGCTAGTGTGCTTAAAGATGGTTGTGATGTTGGTTTTTGTCCACA
-----------------------H-H-I-H-H----------------------------- S612
CTGAAACATTTCATGACACAATTCAATTCACGACAACAGACACGATCACACGAATTCTACCAACACTACAACCAAAAACAGGTGT
C K S T W W E V K S A V V C A S W L K D G C D W G F C P H
Replicase 1 a
CAGACATAAATTGCGTTCACGTGTTAAGTTTGT TAATGGACGTGTTGTTATTACCAATGTTGGTGAACCTATAATTTCACAAccTTCTAAGT
H-H-I------------------------------------------------ 5704
GTCTGATTTAACGCAAGTGCACAATTCAAACAATTACCTGCACAACAATAATGGTTACAACCACTTGGATATTAAAGTGTTGGAAGATTCA
R H K R S R V K F. V N G R V V T N W G E P S C P S K
Replicase 1a
TGCTTAATGGTATTGCTATACAACATTTTCAGGTTCTTTTGATAACGGTCACTATGAGTTTAGATGCTGCTAATAATGCTGTCTATGAT
H-----Hee-i-H----------------------------------H-- 5796
ACGAATTACCATAACGAATATGTTGTAAAAGTCCAAGAAAACTATTGCCAGTGATACATCAAATACTACGACGATTATTACGACAGATACTA
L. L N G : A Y T T. F. S. G. S F D N G H Y V V Y D A A N N A V Y D
Replicase 1 a
GGTGCTCGTTTATTTGCTTCAGATTT ACTTTAGCTGTTACAGCTATTGTGTAGTAGGTGGTTGTGTAACATCTAATGTTCACAAT
-------------------------------------------------------------------- 5888
CCACGAGCAAATAAACGAAGTCTAAACAGATGAAATCGACAATGTCGATAACAACATCATCCACCAACACATTGTAGATTACAAGGTGGTTA
A R L. F A S D L S T L A W T A W W W G G W T - S N W P P
Replicase 1 a
TGTTAGTGAGAAAATTTCTGTTATGGATAAACTTGATACTGGTGCACAAAAATTTTTCCAATTTGGTGATTTTGTTATGAATAACATTGTTC
-------------------asse------------------------------------- iss80
ACAATCACTCTTTT AAAGACAATACCTATTTGAACTATGACCACGTGTTTTTAAAAAGGTTAAACCACTAAAACAATACTTATTGTAACAAG
v S E K I S v M D K L D T GReplicase
A 0 1aK F F O F G D F v M N N i v
TGTTTTTAACTTGGTTGCTTAGATGTTTAGTCTTACGTACTTTATTATGAAGCATGAATAAAGTATTGCCAAGGC TCCTAAACGT
---------------------------------as----------------------------------------------- 6072
ACAAAAATGAACCAAccAACAFACAAATCAGAAAATGCAGAAGATAAFACTCGTACTATAATTTCAATAACGGTCCGAGGATTGCA
F L T W s M F S L L R S J M K H D K V A K A P K R
Raplicase 1a
ACAGGTGT TATTTTGACACGTAGTTTTAAGTATAACATTAGATCTGCTTTGTTTGTTGTAAAGCAGAAGTGGTGTGTTATTGTT ACTTTGTT
-------------------------as-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-----------------------------------ee- 616
TGTCCACAATAAAACTGTGCATCAAAATTCATATTGTAATCTAGACGAAACAAACAACATTTCGTCTTCACCACACAATAACAATGAAACAA
G W L T R S F K Y N R S A F W W K 0 K. W C W W T L F
Replicase 1a
TAAGTTCTTATTGTTAT TATAT ccTATTTATGCAct TGTTTTTATGATTGTGCAATTTAGTCCTTTTAATAGTCTTTTATGTGGTGACATTG
-------------------------------------see------------------r-H----------------or 6256
ATTCAAGAATAACAATAATATACGATAAAT ACGTGAACAAAAATACT AACACGTTAAATCAGGAAAATTATCAGAAAAT ACACCACGTAAC
K F L L L L Y A 1 Y A L V FM.
--or Replicase 1 av D F S P F. N S L L C G D
TAAGGGTTATGAAAAATCC ACTTTTAATAAGGAATTATTGGGTAATTCTATGGTTTGTAAGATGGTTTGTTAGTTACAAGAGTTT
-------------------------------------------all-anal-a------------ee-ee-------------------------- 63.48
ATTCACAATACTTTTTAGGTGAAAAATTCCTATAAAAACACCAFTA AGATACCAAACATTCACACAAACAAATCAATAGTTCTCAAA
W S G Y E K S T F M K o Y C G N S M W C K M C - F S Y O E F
-- Replicase 1 a
AATGATTTGGATCAT ACTAGTCTTGTTTGGAAGCACATTCGTGATCCTATATTAATCAGTTTACAACCATTTGTTATACTTGITATTTTGTT
---------------------------e-r-e----------------------------------H------------------------- 644O
TTACTAAAccTAGTATGATCAGAACAAAccTTcsTGTAAG ACTAGGATATAATAGTCAAAGTGGTAAACAATATGAACAATAAAACAA
N h S w w K H R D P S - O P F W L W
Replicase 1a
AATTTTTGGTAATATGTATTTGCGTTTTGGACTTTTATATTTTGTTGCACAATTTATTAGTACTTTTGGTTCTTTCTTAGGCTTYCATCAGA
-----------------------------------------------es---------------ee-les---------------------- 6532
TTAAAAACCATTATACAAAACGAAAACCTGAAAATATAAAACAACGTGTTAAATAATCATGAAAACCAAGAAAGAATCCGAAAGAGTCT
F G N M Y L R F G L L Y F W A C F S T F G S F L G F H O
Replicase 1 a
AACAGGGTTTTACATTTTTCCGTTTGAGTTTTAAATGAGTTTTTAGCTACATTATTGTCTGcAAAATTGTTTTATTTTTAGA
----------------------------------------ee-eeee--------------------------------- 662
(CACCAAAAAGTAAAACACGGCAAACTACAAAATACATTACT CAAAAATCGATGTAAATAACAGACGTTTTAACAAAATAAACAATC
K 0 w F L H F v P F D v L C Replicase
NEFL A T F 1 v C K 1 v L. F V R
1a
CATATTATTGTTGGCTGTAATAATGCTGACTGTGTAGCTTGTTCTAAAAGTGCTAGACTTAAACGTGTACCACTTCAAACTATTATTAATGG
GTATAATAACAACCGACATTATTACGACTGACACATCGAACAAGATTTTCACGATCTGAATTTGCACATGGTGAAGTTTGATAATAATTACC
H 1 i v Gt C N N A D C v A C S K S A R L. K. R V. P. L. O. T N G
Replicase 1 a
TAGCATAAATCATCATGAAGCAAGGGGTACTTGTTCTsAAAAACATAACTTCTTTGTGTTAATTGTGATTCTTGGGC
------------------------------as-s-s-s---------------------------------------------------------- 6808
ATACGTATTTAGTAAGATACAATACGATTACCACCATGAACAAAGACATTATTTGTATTGAAGAAAACACAATAACACTAAGAAAACCCG
M H K S F Y V N A N G G T C F C N K H N F F C v N C D s f G
-Replicase a
CTGTAAAcTTTTATTAATGGTATATTGCAAGAGAGCTTGGTAATGTTGTTAAAACAGCTGTTCAACCCACAGCTCCTGCATAGTAT
-----------------------------------------He--------------------------------------------------- 6900
GACCATATGAAAATAATTACCACTATAACGTCCTCGAACCATTACAACAATTTTTCGACAAGGGGTGTCGAGACGTATACAATAA
P G N T F N G D A R E L G N V V K T A V O P T A P A Y V
Replicase 1a
ATTGATAAGGTAGATTTTGT TAATGGATTTTATCGTCTTTATAGTGGTGACACTTTTTGGCGGTATGACTTTGACATTACTGAATCTAAGTA
---------------------------------------------------------------------------------------------- 6992
TAACTATTCCATCTAAAACAATTACCTAAAAT AGCAGAAATATCACCACTGTGAAAAACCGCCATACTGAAACTGTAATGACTTAGATTCAT
O K W o F W N G F Y R Y S G D F W R Y o F .D T E S K Y
Replicase 1a
TAGTTGTAAAGAGGTTCTGAAGAATTGTAATGTTTTAGAAAATTTTATTGTTTACAATAATAGTGGTAGTAACATTACACAGATTAAAAATG
------------------------------------------------------------------------- 7084
ATCAAcATTTCTCCAAGACTTCTTAACATTACAAAATCTTTTAAAATAACAAATGTTATTATCACCATCATTGTAATGTGTCTAATTTTTAC
S C K E W L K M C N W L E N F W Y N N S G S N T O K N
Replicase 1 a
CIGTGTTTATTTTTCTCAA GTGGTGAACTATAAAGGGTAAATTCAGAGTTGTGTCAACTTTATAGTTGATTTAATGGTGTT
-------------------------------------------as-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s 7 78
GAACACAAATAAAAAGAGTTAACAACACACTTGGATATTTCAACCATTTAAGTCTCAACAACAGTTGAAATAGTCAACTAAAATTACCACAA
A C v Y F s O L L C E P K L V N S E L L S T L S v D F N G v
Replicase 1a
TTGCATAAGGCATAreTTGATGTTTTGTGTAATAGTTTTTTTAAGGAGCTAACTGCTAACATGTCCATGGCTGAATGTAAAGC ACACTTG
----------------------------asse------a-ha-ma-e-less i268
AACGATTcCGTATACAACTAcAAAACACAATCAAAAAAAICCTCGATGACGATTGTACAGGTACCGACTTACATTTCGATGTGAACC
H K A Y V D v L. C N S F F K E L T A N M S M A E C K A T L G
Replicase 1a m
TTTGACTGTTTCTGATGATGATTTTGTTTCAGCTGTTGCCAATGCACATAGGYATGACGTTTTGCTTTCAGATTTGTCATTTAATAATTTTT
--------------------------------------------------- 7360
AAACTGAAAAGACTACT ACTAAAACAAAGTCGACAAGGTTACGTGTATCCATACTGCAAAACGAAAGCTAAACAGTAAATTATTAAAAA
T V S D D D F W S A v A N A H R Y D V L L S D L S F N N F
Replicase 1 a
TATTTCYTATGCTAAACCTGAAGATAAGTTGTCCGTTTATGACATIGCTTGTGTATGCGTGCCGGTTCTAAGGTTGTAACCATAATGTT
-------------------------------------------------------------------------------------- 752
AATAAAGAAT AGATTGGAct TCTATAACAGGCAAATACTGTAACGAACAAC ATACGCACGGCCAAGATTCCAACAATTGGTATTACAA
F S Y A K P E D K L S W Y D A C C M R A G, S K V V N H N W
Replicase 1 a
k
TTAATCAAAGAGT CAATACCTATTGTTTGGGGTGTCAAGGACTTAATACTCTTTCTCAAGAAGGTAAGAAGTACCTTGTAAAACAACAA
AATTAGTTTCTCAGTTAGGATAACAAACCCCACAGTTCCTGAAATTATGAGAAAGAGTCTTCATTCTTCATGAACAATTTTTGATT
L K E S 1 P : V w G V K D F N T L S 0 E G K K Y L. v K T T K
Replicase 1 a m
AGCAA AGGGTTTGACTTTTTTATAACTTAATGATAACCAAGCAATTACACAAGCCTGCACAGTATATTGAAAACAGGGGCTG
------------------------------ee-us-------------------------------------------- 76.36
CGTTTCCCAAACTGAAAAAATAATTGAAAATTACTATTGGTTCGTAATGTGTCAAGGACGATGATCATATCAACGTTTTGTCCCA CGAC
A K G L T F L L T F N D N A O W P A S W. A. K. O. G. A
Replicase 1 a
GTTTTAAACGT ACTTATAATTTTCGTGGTATGTATGTTTATTGTTGTTGCATTGTTTATTGGTGTCTCATTTATTGATTATACAACCACT
H--------------------------------------------------------------------------------------- 7728
cAAAATTTGCATGAAT ATTAAAAGACACCATACATACAAATAAACAACAACGTAACAAATAACCACAGAGTAAATAACTAATATGTTGGTGA
G F K R T Y N F L W Y V C L. F V V A. L. F I G V S F | D Y T T Y
Replicass 1a
GTAACTAGCTTTCATGGTTATGATTTTAAGTACATTGAGAATGGTCAGTTGAAGGTGTTTGAAGCACCTTT ACACTGTGTTCGTAATGTTTT
---ee-le-ele-------------------------------------------------------------- 782O
CATTGATCGAAA(TACCAAACAAAATTCATGT AACTCTTACCAGTCAA CTCCACAAACTTCGTGGAAATGTGACACAAGCATTACAAAA
W T S f H G Y D F K Y E N G O L K V F E A P C W R N V F
Replicase 1 a
TGATAATTTTAAT CAATGGCAGAGGCTAAGTTTGGTGTTGT TACT ACTAAT AGTGATAAATGTCCTATAGTTGTTGGTGTTTCAGAGCGTA
ACTATTAAAATTAGTT ACCGT ACTCCGATTCAAACCACAACAATGATGATTATCACTATTTACAGGATATCAACAACCACAAAGTCTCGCAT
D N F N. O. w H E A K F G V v T T N S D K C P. V V G V S E R
Replicase 1a- - -
TTAATGTTGTTCCTGGTGTTCCAACAAATGTATATTTGGTAGGAAAGACTCTTGTTTTTACATTACAGGCTGCTTTTGGAAACACAGGTGTT
AATTACAACAAGGACCACAAGGTTGTTTACATATAAACCATCCTTTCTGAGAACAAAAATGTAATGTCCGACGAAAACCTTTGTGTCCACAA
N V V P G W P T N V Y L. W G K W F T L O A A F G N T G V
Replicase 1a
TGT TATGACTTTGATGGTGTTACCACTAGTGATAAGTGTATTTTTAATTCTGCTTGT ACTAGGTTGGAAGGTTTGGGTGGTGACAATGTTTA
ACAATACGAAACACCACAATGACACTATCACAAAAAATTAAGACAACAGATCCAAC CTCCAAACCCACCACGTTACAAAT
C Y D F D G v T T S D K C F N S A C T R L E G L G G D N V Y
Replicase 1a
TTGTTACAAcACTGATCTTATTGAAGGTTCTAAACCTTATAGTATTTTACAGCCCAATGCTTATTATAAGTATGATGTTAAAAATTATGTAC
--------------------------------------------------------------------------------------- 888
AACAATGTTGTGACTAGAATAACTTCCAAGATTTGGAATATCATAAAATGTCGGGTTACGAATAAATTCATACTACAATTTTT AATACATG
C Y N T D L I E G S K P Y S O P N A Y Y K Y D V K N Y W
Replicase 1a
GTTTTCCAGAAATTTTAGCT AGAGGTTTTGGCTTACGT ACTATTAGAACTTTGGCTACACGTTATTGT AGAGTTGGTGAATGCCGTGACTCA
CAAAAGGTCTTTAAAATCGATCTCCAAAACCGAATGCATGATAATCTTGAAACCGATGTGCAATAACATCCAACCACTTACGGCACTGAGT
R F P E L A R G F G L R T R T L A T R Y C R W G E C R D S
--Replicase 1 a
CATAAAGGTGTTT GTTTTGGTTTTGATAAATGGTATGTTAATGATGGACGTGTTGATGACGGTTACATTT GTGGTGATGGTCTTATAGACCT

-------H-----------------her--------------------------------------- 8372
GTATTTCCACAAACAAAACCAAAACTATTT ACC ATACAATTACTACCTGCACAACTACTGCCAATGTAAACACCACTACCAGAATATCTGGA
H K G V C F G F D K W Y V NDG R v D D G Y
Replicase 1a C G D G L D. L.
TCTGTTAATGTACTCTCAATCTTTAGTTCATCTTTTAGCGTTGTGGCTAGTCGGACATATGTTGTTTAATTTCTTTTTGCAGCATTA
----------------------ee-H---H-------------------H----- 8.464
AGAACAATTACATGAGAGTTAGAAATCAAGTAGAAAATCGCA ACACCGATACAGACCTGTATACAACAAATAAAAGAAAAACGTCGTAAAT
V N W L S F S S S F S W W A M S G H M L F N F L F A A F
Replicase 1a
TTACATTTTTGTGCTTTTTAGTTACTAAATTTAAACGTGTTTTTGGTGATCTTTCTTATGGTGTTTTTACTGTTGTTTGTGCAACTTTGATT
H-------------H-e-H----------Hees-H--------H--------- 8556
AATGTAAAAACACGAAAAATCAATGATTTAAATTTGCACAAAAACCACTAGAAAGAATACCACAAAAATGACAACAAACACGTTGAAACTAA
T F c F W T K F K R V F G O L S Y G V F T V V C A T L
Replicase 1 a
AAAACAt TTCTTATGTTGTTACT CAAAATTTATTTTTTATGTTGCTTTATGTATTTTGTATTTTGTTTTTACTAGGACAGTGCGTTATGC
TATTGTAAAGAATACAACAATGAGTTTTAAATAAAAAATACAACGAAATACGATAAAACATAAAACAAAAATGATCCTGTCACGCAATACG
N N S Y v v T. O. N. L. F F M L L Y A L Y F W F T. R T V R Y A
Replicase 1a
TTGGATTT GGCATATTGCATACATTGTTGCATACTTCTTGT TAATAccATCGTGGCTTCT CACATGGTTTAGTTTTCCTGCATTTTTAGAGC
H-----------------H--------------------------------------- 8740.
AACCTAAACCGTATAACGTATGTAACAAC GTATGAAGAACAATTATGGACCACCGAAGAGGTACCAAATCAAAACGACGTAAAAATCTCG
w I W H A Y i v A. Y F L L 1 P W W L L T W F S F A A F E
Replicase 1a- -
TTTTACCTAATGTTTTTAAGTTAAAAATCTCTACT CAATTGTTTGAAGGTGATAAGTTTATAGGTACTTTTGAGAGTGCTGCTGCAGGTACA
a---------------------all----------------------------------H------------ 88.32
AAAAGGATTACAAAAATTCAATTTTTAGAGATGAGTTAACAAACTTCCACATTCAAATATCCAGAAAACTCTCACGACGACGTCCATGT
L. L. P N V F K L K S T O L. F E G D K F G T F E S A A A G T
--Replicase 1 a
TTTGTTCTTGACATGCGTTCTTATGAAAGGCTGATAAATACTATTTCACCTGAGAAACTTAAGAATTAfGCTGCAAGTTATAATAAATATAA
-----------------------------------H---------------------- 8924
AAACAAGAACTGTACGCAAGAATACTTTCCGACTATATGATAAAGTGGACTCTTTGAATTCTAATACGACGTCAATATATTTATAT
F W L D M R S Y E R L N T S P E K. L. K. N. Y. A A S Y N K Y K
-Replicase 1 a
At ATTATAGTGGTAGTGCTAGTGAGGCTGATTATCGTTGTGCTTGTTATGCTCATTTAGCCAAGGCTATGTTAGATTACGCAAAAGATCATA
----------------------------H------------------ 906
TATAATAT CACCATCACGAT CACTCCGACTAATAGCAACACGAACAATACGAGTAAATCGGTTCCGATACAATCTAATGCGTTTTCTAGTAT
Y Y S G S A S E A D Y R C A C Y. A H. L. A K A M L D Y A K D H
Replicase a
ATGACATGT TATATTCTCCACCTAccATTAGCTACAATTCCACCTTACAATCTGGTCTTAAGAAGATGGCACAACCATCTGGTTGTGTTGAG
H-H----------------------H----------------------------- 9 OB
TACTGTACAATATAAGAGGTGGATGGTAATCGATGTAAGGTGGAATGTTAGACCAGAATTCTTCTACCGTGTTGGTAGACCAAcACAAcTC
N D M Y S P P T S Y N S T O S G K K M A C P S G C W E
Replicase 1a
AGATGTGTGGTTCGCGTCTGTTATGGTAGTACTGTGCTTAATGGAGTTTGGTTAGGTGACACTGT TACTTGTCCTAGACATGTCATAGCAcc
------------------------------------------------------------estro-------------------------- g2OO
TCACACACCAAGCGCAGACAAt ACCATCATGACAcGAATTACCCAAACAATCCACGTGACAATGAACAGGACGTACAGTAtCGTGG
R C V V R W C Y G S T W - N G W W L G D T W T C P R H V A P
-Replicase 1a m
ACAACCACGCTATTGATTATGACAGCATAAGTACTATGcGTTTGCATAATTTTCAGTGTCCATAATGGTTCTTCTTGGGAG
-------------------------------------------------------------------------------------- 9292
AGTTGGTGACAAGAATAACTAATACTAGTACGTATATCATGATACGCAAACGTATTAAAAAGTCACAGAGTATTACCACAGAAGAACCCTC
S T T W D Y D H A Y S T M R L H N F S W S h N G V F L G
Replicase 1 a
TTTTGGTGTTACAATGCATGGTTCTGGTTGCGATAAGGTTTCACAATCTAATGTACATACACCTAAACATGTTTTTAAAACGTTGAAA
--------------------------------------------------------------------------------------ee------ 938.
AACAACCACAATGTACGTACCAAGACACAAceCAAATCCAAAGTGTAGATTACATGATGTGGATTTGACAAAAATTTTGCAACTT
W W G W T M H G S W L R K V S O S N W H T P K H. W. F. K T L K
Replicase 1a
CCTGGGCTCTTTTAA, ATTTTAGCATGTTATGAAGGTATTGCATCTGGTGTTTTTGGTGTTAATTTACGTACAAACTTTACT ATTAAAGG
---------------------------ee-les-s-s-s-s-s-s-----------ee-as-s-s-s-s-s-s-rele---is 976
GGACCACGAAGAAAATTATAAAATCGTACAAT ACTTCCATAACG TAGACCACAAAAACCACAATTAAATGCATGTTTGAAATGATAATTTCC
P G A S F N L. A C Y E G A S G V F G V N L R T N F T K G
Replicase 1a--
TTCTATAAAGGACTTGGGTTCTCCTGGTTATAATGTTAGAAATGATGGTACTGTTGAGTTTGTTATTACACCAAATTGAGTAG
AAGAAAATATTACCTCGAACACCAAGAGGACCAAATTACAATCTTTACT ACCAGAAACTCAAAACAATAAATGTGGTTTAACT CAATC
S F N G A C G S P G Y N V R N D G T :V E F C Y L H O E L
-Replicase 1 a A
GT AGTGGGCTCATGTTGGTTCTGATTTT ACTGGTAGTGTTTATGGTAATTTTGATGACCAACC TAGTTTGCAAGTTGAGAGTGCCAACCTT

------------a-He-as-s-s-s-s-s-s-s-s-s-s----------------------------------------- 966O
CAT CACCACGAGTACAACCAAGACT AAAATGACCATCACAAATACCATTAAAACTACTGGTTGGATCAAACGTTCAACTCTCACGGTTGGAA
G. S S A H W G S D F G. S. V Y G N F D d O P S L O W E S A N t
Replicase 1a
ATGCTATCAATAATGGCCTTGATGCGCTTGTTGAATGGTTGTAGGTGGTGGTGGTTCAACTAGACTTAATGTTGATG
TACGAAGTCATACAACAAGGAAAAACATACGACGAAACAACTACCAACACCACCACCAACGCAAGTTGATCCAATTACAACTACC
L S N W W A F - Y A. A N G C R W W R S T R V N V to G
Replicase 1a
TTTTAATGAATGGGCTATGGCTAATGGTTATACAATTGTTTCTAGTGTTGAGTGCTATTCTATTTTGGCAGCAAAAACTGGTGTTAGTGTTG
s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s------- 984
AAAATTACTTACCGAACCGATACCAATAGTAACAAAGACACAACTCACGATAAGAAAAACCGTCGTTTTGACCACAATCACAAC
F N E W A M A N G Y T W S S W E C Y S . L. A. A K T G V S V
Replicase 1 a
AAAAGTATTCATTCAA CATCFTCATGAAGGTTTGGTGGTAAAAACATACTTGGTTATTCTAGTTATGTGAGAGTTCACACTA
-----------------------------------eeeee----------------------------- 9936
TTGAACAATCGAAGGTAAGTGAGAAGTACCCAAAACCACCATTGATGAACAATAAGATCAAAT ACACTACCAAGTGTGA
E. O. L. L. A S 1 O H L H E G F G G. K. N. l G Y S S C D E F T L
- Replicase 1a
GCTGAAGTTGTGAAGCAGATGTATGGTGTTAACTTGCAAAGTGGTAAGGTTATTTTTGGTTTAAAAACAATGTTTTTATTTAGcGTTTTCTT OO28
CGACTTCAACACTTCGTCTACATACCACAATTGAACGTTTCACCATTCCAATAAAAACCAAATTTTTGTTACAAAAATAAATCGCAAAAGAA
A E V V K Q M Y G V N L O S G. K. v . F G L K T M F L F S v F F
Replicase 1a
CACAATGTTTTGGGCAGAACTCTTTATTTATACAAACACTATATGGATAAACCCTGT TATACTTACACCTATATTTTGTTT ACTTTGTTTT
---------------------H-----------------------------H------------------- O2O
GTGTTACAAAACCCGTCTTGAGAAATAAATATGTTTGTGATATACCTATTTGGGACAATATGAATGGGATAAAAACAAATGAAAACAAAA
T M F W A E L F Y T N T W N P V 1 T P F C - L. L. F
Replicase 1a
TGTCATTAGTTTTAACTAGTFTCTTAAACATAAGTTTTTGTTTTTGCAAGTATTTTTATTACC TACTGTTATGcAACTGCTTTATATAAT
-H--------------------------He------------ 10212
ACAGTAATCAAAATTGATACAAAGAATTTGTACAAAAACAAAAACGTCATAAAAATAATGGATGACAATAACGTTGACGAAATATATTA
S L W L T M F K H K F - F - O W, F L P T V A A Y N
Replicase a -
GTGTTTTGGATTATTAAT AGAAAATTTTTGGCTGACCATTTTAACTATAATGTTTCAGTATTACAAATGGATGTTCAGGGTTTAGTTAA
-------H-----Hors---------------------------------------------------- O3Ol
ACACAAAACCTAATAATGTATCATTTTAAAAACCGAct GGTAAAATTGATATTACAAAGTCATAATGTTTACCTACAAGTCCCAAATCAATT
C. W. L. D Y Y V K F L A D H F N Y N V S V O M D V O G L V N
Replicase 1 a
TGTTTTGGTCTGTTTATTTGTTGTATTTTTACACACATGGCGTTTTTCTAAAGAACGTTTCACACATTGGTTTACATATGTGTGTTCTCTTA
--------H-----------------------------H-------------------------- 10396
ACAAAACCAGACAAATAAACAACATAAAAATGTGTGTACCGCAAAAAGATTTCTTGCAAAGTGTGTAACCAAATGTATACACACAAGAGAAT
v L. v c L F v v F L H T W R F S K E R F T H W F T Y V C S L
Replicass 1a-H
TAGCAGTTGct TACACTATTTTTATAGTGGTGACTTTTTGAGTTTGCTTGTTATGTTTTTATGGCTATATCTAGTGATTGGTACA, TGGT
o-------------------------------------------Ho-Ho- Ot.88
ATCGTCAACGAATGTGAATAAAAAATCACCACTGAAAAACCAAACGAACAATACAAAAAACACGATATAGATCACTAACCATGTAACCA
A W A Y T Y F Y S G D F L S W. M. F. C A S S D W Y G
m Replicase 1a
GCCATTGTTTTTAGGTTGTCACGTTTGATTATATTTTTTTCACCTGAAAGTGTATTTAGTGTTTTTGGTGATGTGAAAcTCACTTTAGTTGT
-------------------------------------------------------H-I-H O580
CGGAACAAAAATCCAACAGTGCAAACTAAATAAAAAAAGTGGACTTTCACATAAATCACAAAAACCACTACACTTGAGTGAAATCAACA
A v F R L S R L F F S P E S v F S v F G D v K L T L V v
Replicase 1 a
TATTAATTTGTGGTTATATTTGACTTATTGGGGCATTGATTGGTTCAAAGGTTTTTTAAATGACTATGGGTGTTTATGATT
----------------------------------------------------------------- Os2
AATAAATTAAACACCAATAAATCAAACATGAATAACCCCGTAAAACAAACCAAGTTATCCAAAAAATTACAGATACCCACAAATACTAA
Y C G Y L V C T Y. W G Y W F N R F F K C T M G V Y D
Replicase 1 a
TTAAGGTGAGTGCTGCTGAATTTAAATACATGGTTGCTAATGGACTTCATGCACCATATGGACCTTTTGATGCACTTTGGTTATCATTCAAA
p-H-----------------------Hero-e-Ha-e-------------------------- 1076.
AATTCCACTCACGACGACTTAAATTTATGTACCAACGATTACCTGAAGTACGTGGTATACCTGGAAAACTACGTGAAACCAA TAGTAAGTTT
F K W S A A E F K Y M W A N G L H A P Y G P F O A W L S F K
-Replicase 1a
TT ACTTGGTATTGGTGGTGACCGTTGTATAAAAATTCAACGTCCAATCCAAACTGACTGATTGAAGTGTACTAATGTTGTGTATTGGG
------es---------------------------------------------------------------------- O856
AATGAACCAAACCACCACTGGCAAAT ATTTAAAGTTGACAGGTTAGGTGACTGACAAACTTACATGATTACAACACAATAACCC
L G G G D R C 1 K S T V C S K L T D L K C T N V V L. L. G
Replicase a f
TTGTTTGTCTAGTATGAACATTGCAGCTAATTCTAGTGAATGGGCTTATTGTGTTGATTT ACACAATAAGATTAATCTTTGTGATGACCCAG
H------------------------------------------H--- O948
AACAAACAGATCATACTTGTAACGTCGATTAAGATCACACCCGAATAACACAACTAAA GTGTATTCAATTAGAAACACTACTGGGC
C L S S M N A A N S S E W A Y C W D H. N. K N D. D. P
Replicase 1 a
AAAAAGCTCAAGGTATGTTGTTAGCACTCCTTGcGTTCTTTCTAAGTAAACATAGTGATTTTGGTCTTGATGGCCTTATTGATTCTTATTTT
--------------------------------------------------------------------- 1040
TTTTTCGAGTTCCATACAACAATCGTGAGGAACGCAAGAAAGATTCATTTGTATCACTAAAACCAGAACTACCGGAATAAC TAAGAATAAAA'
E. K. A G M A A F F S K H S D F G L D G L E D S Y F
Replicase 1 a
GATAATAGTAGCACCCTGCAGAGTGTTGCTTCATCATTTGTTAGTATGCCATCATATATTGCTTATGAAAATGCTAGACAAGCTTATGAGGA
-------------------------------------s-s-s-s-erer--ee----------------------Hire---------------------- 1:32
CTATATCATCGGGACGCTCACA ACGAATAGTAAACAATCATACGTAGTATATAACGAATACTTTACGATCTGTCGAAT ACTCCT
D N S S T L Q S v A S S F W S M P S Y A Y E N A R O A Y E
Replicase ta
TGCTATTGCTAATGGATCTTCTTCTCAACTTATTAAACAATTGAAGCGTGCCATGAAT ATCGCAAAGTCTGAATTGATCATGAGATATCTG
------------------------------------as-s-s-s-s-areasons-eless-as-as-s-s-s-see-testernessee 1224
ACGATAACGATT ACC TAGAAGAAGAGTTGAATAATTTGTTAACTTCGCACGGT ACTTATAGCGTTTCAGACTTAAACTAGTACTCTATAGAC
A A N G S S S O K K R A M N A K S E F D E S
Replicase 1 a
TTCAGAAGAAAATTAAT AGAATGGCTGAACAAGCTGCTACCAGATGTAAAAGAAGCACGCTCTGTTAATAAAAATCTAAAGTTAFTAGT
-------ee-------------------------------------arole-la----ee-ee-eeeeeee-eel-as-s-s- 1316
AAGCCTTTTAATTACTTACCGACTGTTCGACGATGAGTCACATA, TCTCGGCGAGACAATTATCTTAGATTTCAATAATCA
W 0 K K N. R. M. A E G A A T O M Y K E A R S V N R K S K W S
Replicase 1 a
CTATGCACTCTTACTTGGAATGTTAAGACGTTTGGATAGTCTAGGGAAACTGTTTGAATTAGCACGTGATGGTGTTGTGCC
re -------------------------------------------------------------------- 1:03
CGATACGTGAGAAATGAAAAACCTTACAATTCTGCAAACCTATACAGATCACAACTTTGACAAAACTTAAATCGTGCACTACCACAACACGG
A M H S L F. G M L R R D M S S W E T W L N - A R D G V V P
Replicase 1a
A TGTCAGTTATACCGC AACTTCAGCTTCCAAACTAACTATTGTTAGTCCAGAT CTTGAATCTTATTCTAAGATTGTTTGTGATGGTTCG
re-es---as-as-a- ---------------------------------ee-ee---------------------------- SOO
AAACAATAGGACGTTGAATCGAAGGTTTGATTGAAACAATCAGGTTAGAACTTAGAATAAGATTCA ACAAACACTACCAAGAC
L S v i P A T S A S K L T i v S P D L E S Y S K i v C D G S
Replicase 1a
TTCATTAGCTGGAGTTGTTTGGACACTTAATGATGTTAAAGACAATGATGGTAGACCTGTTCATGTTAAAGAGATTACAAGGGAGAATGTT
AAGAAACGACCTAACAAACCGTGAATTACTACAATTCGTTACACCATCGGACAAGTACAATTTCTCTAATGTTCCCTCTTACAA
V :H Y A G V V W T L N D V K D N D G R P W H W" K E T R E N V
-Replicase 1 a
GAAACTTTGACATGGCCTCTTATccT TAATTGTGAACGTGTTGTTAAACTTCAAAATAATGAAATTATGCCTGGTAAACTTAAGCAAAAACC
-H---------------------------------------------------------- 6Bq.
CTTTGAAACT G T ACCGGAGAATAGGAATTAACACTTGCACAACAATTTGAAGTTTTATTACTTTAATACGGACCATTTGAATTCGTTTTTGG
E T L T W P L I L N C E R V Replicase
V K 1a O N N E 1 M P G K L K Q K P
TATGAAAGCTGAGGGTGATGGTGGGTTTTAGGTGATGGTAATGCTTGTATAATACTGAGGGTGGTAAAACTTTATGTATGCTTATATTT
-------------------------Hoo-o-o-H----------H-----H--- 1776
ATACTTTCGACT CCCACTACCACCACAAAATCCACTACCATTACGAAACATATTATGACTCCCACCATTTTGAAAATACATACGAATATAAA
M. K. A E G D G G V G D G N A Y N T. E. G. G. K. F M Y A Y
Replicase 1 a
CTAATAAAGCTGACCTTAAATTGTTAAGTGGGAGTATGAGGGGGTTGCA ACACAATCGAGTTAGACTCTCCTTGTCGATTATGGTCGAA
-------------------------------------------------------- 1868
GATTATTTCGACTGGAATTTAAACAATTCACCCTCATACTCCCACCAACGTTGTGTAGCTCAACTGAGAGGAACAGCTAAATACCAGCTT
S N. K. A D K F W K W E Y E G G C N T E L D S P C R F M W E
Replicase 1a
ACACCTAATGGTCCTCAAGTGAAGTATTTGTATTTTGTTAAAAATAAATACCTTAC GTAGAGGTGCCGTTCTTGGTTTTATAGGTGCCAC
------------------------------------------------H---------------- 1960
TGTGGATTACCAGGAGTTCACTTCATAAACATAAAACAATTTTTAAATTTATGGAATGCATCTCCACGGCAAGAACCAAAATATCCACGGTG
T P N G P o V K Y Y F W K N - N T L R R G A W L G F I G A f
Replicase 1 a
AATCGTC TACAAGCTGGTAAACAAACTGAATTGGCTGTTAATTCTGGACTTTTAACTGCTTGTGCTTTTTCTGTTGATCCAGCAACCACTT
TAAGCAGATGTTCGACCATTGTTGACTTAACCGACAATTAAGAccGAAAATTGACGAACACGAAAAAGACAACTAGGTCGTTGGTGAA
R L. O A G K T E L A V N S G L L T A C A F S W D P A T T
Replicase a
ACTTGGAAGCTGTTAAACATGGTGCAAAACCTGTAAGTAATTGTATTAAGATGT TATCTAATGGTGCTGGTAATGGTCAAGCTATAACAACT
--------------------------------------------------------------------------------- 2144
GAACCTTCGACAATTTGTACCACGTTTTGGACATTCAFTAACATAATTCTACAAT AGATTACCACGACCATACCAGTTCGATATTGTTGA
Y L. E A V K H G A K P W S N C K M L S N G A G N G D A T
Replicase 1a
AGTGTAGATGCTAACACCAATCAAGATTCTTATGGTGGAGCGTCTATTTGTTTGTATTGTCGGGCCCACGTTCCTCACCCTAGTATGGATGG
-------------------------------------------------------------------- 2236
TCACAFCTACGATTGTGGTTAGTTCTAAGAATACCACCTCGCAGATAAACAAACATAACAGCCCGGGTGCAAGGAGTGGGATCATACCTACC
S W D A N T N C D S Y G G A S C L Y C R A H W P H P S M D G
Replicase 1a
TTACTGTAAGTTTAAGGGTAAATGTGTTCAGGTTCCTATTGGTGTTGGATCCTATAGGTTTGAGAAAATAAGGTGTAATGTTT
--Ha-e------------------------em-Hees------real-e-Ha----------------------------- 12328
AATGACATTCAAATTCCCATTTACACAAGTCCAAGGATAACCAACAAACCTAGGATAATCCAAAACAAATCTTATTACACACATTACAAA
Y C K F. K. G. K. C. W. O. W P G C D P J R F C E N N W C N V
Replicase 1a
GTGGTTGTTGGGGGACACGGGGTGCTTGTGATCGACAACCATTCAAAGGTTGACATTTCTTATTTAAAcGAGCAAGGGGTTCTAGTG
------ee---------------s-s-s-s-s-s-s-s-s-s-s-s--es--------------------------------- 22O
CACCAACAACCAACCCTGTGCCCACACGAACACTAGCATGTTGGTAAGTTTCACAACTGT AAAGAATAAATTTGCTCGTTCCCCAAGAT CAC
C G C W L G H G C A C D R T T O S v D S Y N E O G v L v
Replicase 1a
R A R G S S
Replicase is
CACCGACTAGAACCCTGTAATGGCACGGACATCATAAGTGTGTCGTGCTTGACATTTATAAAAAAATest TCATCTTGGGTAA
-------------------------arras-a-s-s-s-s-s-s-s-s-at-e-a-s-s-s-s-s-s-s-s-s-s-Hausa---- 12512
TCGACTGATCTTGACATTACCGTGCCGTAGCTATTCACACAAGCACGAAAACTGTAAATATTATTTTTACAAAGTAAGAACCCATTC
O D .
-Replicase 1a
A. A R L. E P C N G O K C W R A F D Y N K N V S F G K
Replicase 1b - -
GTGAAGATGAACTGGTTCGTTTTAAAAATGcGATCTAAGATGGTATTTTGTAAAAGAGGTGTACAAGTCGGrf ATGGAACA
ACAAA CTCTACTTGACACAAGCAAAATTTTAGATAGAATTCCTAccAAAAAACAAATTCTCCACATGATTCAGCCAAACCTTG
C. L. K M N C W R F K. N. A D . K D G Y F W K R C T K S W M E
Replicase 1b.
CCACAATCCAGATAACCAC, AACTTTTCTGGTGCTTGGCTGAGCAGATTCTTACTGGAAAGAGGCAGAGTATTTATGGTA
-------------------- 12696
GCCTTAGGTACAATTGGATGAATTGAAAAGACCACGAAA CGACCACTAAAGAAAGAACTTACCGTCTCAGAAAACA
E O S M Y N L. N. F. S. G A A E H F F T W K D G R W Y G
Replicase 1b
ATGTAG TAGACATAATCT ACTAAATATACTATGATGGACTGGTTAGCTATGCGTAACTTGATGAACAAAATGGATGTTCTAAAA
----------------------------------------------------------------------- 12788
TACAATCATCTGTATTAGAATGATTTATAGATACTACCTGAACCAAATACGATACGCATTGAAACTACTGTTTTAACACTACAAGATT
N V S R H N L T K Y M D L V Y A M R N F D E O N C W K
-Replicase 1b
GAAGTATTAGTTTTAACTGGTTGTTGTGACAATTCTTATTTTGATAGTAAGGGTTGGTATGACCCAGTTGAAAATGAAGATATACATAGAGT
---es-Heer-----------------------reles------------------per------------------------------------ 1288O
CTTCATAATCAAAATTGACCAACAACACTGTTAAGAATAAAACTATCATCCCAACCATACTGGGTCAACTTACTATATGTATCCA
E W L W L T G C C C N S Y F O S K G W Y O P W E N E D I H R W
Replicase 1b.
TTATGCATCTCTTGGCAAAATTGTAGCT AGAGCTATGCTTAAATGCGTTGCTCTATGTGATGCGATGGTTGCTAAAGGTGTTGTTGGTGTTT
AATAC GT AGAGAACCGTTTTAACATCGATCTCGATACGAATTTACGCA ACGAGATACACT ACGCTACCAACGATTTCCACAACAACCACAAA
Y A S L G K i v A R A M L K C v A L C D A M v A K G V V G V
-e-Replicase b
AACATTAGAT AACCAAGATCTTAATGGTAACTTTTAGATTTTGGTGATTTTGTTGTTAGCTTACCTAATATGGGTGTTCCCTGTTGTACA
---------------ress------------ee-eeee-ear-releases. 3O84
ATTGTAATCTATTGGTTCTAGAATTACCATTGAAAATACTAAAACCACTAAAACAACAATCGAATGGATTATACCCACAAGGGACAACATGT
T L D N O D L N G N F Y D F G D F V V S L P N M G W P C C T
-Replicase 1b
TCATATTATTCTTATATGATGCCTATTATGGGTTTAACTAATTGTTTAGCTAGTGAGTGTTTTGTCAAGAGTGATATTTTTGGTAGTGATTT
---He-------------------------------H------------- 3:56
AGATAATAAGAAAACTACGGATAATACCCAAATTGATTAACAAATCGATCACTCACAAAACAGTTCTCACTATAAAAACCATCAct AAA
S Y. Y S Y M M P 1 M G L T N C L A S E C F V K S D F G s D F
Replicase 1b
TAAAACTTTTGATTT GCTTAAGTATGATTTCACTGAACATAAAGAAAATTTATTCAATAAGTACTTTAAGCATTGGAGTTTTGATTATCATC
H-------H----------------------------------------------------- 1328
ATTTTGAAAACTAAACGAATTCATACTAAAGTGACTTGTATTTCTTTTAAATAAGTTATTCATGAAATTCGTAACCT CAAAACTAATAGTAG
K F D L. L. K. Y. D F T E H K E N F N K Y F K - W S F D Y H
-Replicase 1b.
CTAATTGTAGTGACTGTTATGATGATATGTGTGTTATACATTGTGCTAATTTTAATACACTATTTGCCACAACTATACCAGGTACTGCTTTT
-----------------H-------------H-I-H------------------a---------- 13340
GATTAACATCACTGACAAT ACTACTATACACACAATATGTAACACGATT AAAATTATGTGATAAACGGTGTTGATATGGTCCATGACGAAAA
P N C S D C Y O D M C W. h C A N F N T - F A T T P G T A F
Replicase 1b
GGTCCACTATGTCGTAAAGTTTTTATAGATGGTGTTCCACTTGTTACAACTGCTGGTTATCATTTTAAGCAATTAGGTTTGGTTTGGAATAA
------------------He-Ha-H-----------------H---------------------------- 134.32
CCAGGTGATACAGCATTCAAAAATATCTACCACAAGGTGAACAATGTTGACGACCAATAGTAAAATTCGTTAATCCAAACCAAACCTTATT
G P L C R K V F D G W P L V T A G Y H F K C L G u W. W. N. K
Replicase 1b
AGATTTAACACACACTCACTTAGGTTGACAATCACTGAACTTGCAATTGTTACTGACCCTTCCTTGATAATAGCTTCTTCTCCAGCAC
-------------------------------His-Hees-Heerer-H 13524
TCTACAATTGTGTGTGAGTCAATCCAACTGTTAGTGACTTGAAAACGTTAAACAATGACTGGGAAGGAACTATTATCGAAGAAGAGGTCGTG
D V N T H S v R L T | T E ReplicaseOFb-e-Hat
v T D P S L I A S S P A
CGTTGATCAACCACTATTTGTTTTTCTGGCAGCATGAGTACTGGTTTGACAAATCAAGTGTAAGCCAGGTCATTTAATGAAGAG
H---- -H------------------------------------------ 1366
AGCAACTAGTGCGGATAAACAAAAAGAAACGTCGAACTCATGACCAAACTGTTTAGTTCAACAATTCGGTCCAGAAAATTACTTCTC
L V D O R T 1 C F S v A A L S T G L T N 0 V V K P G H F N E E
-Replicase 1b.
TTTTATAACTTCTTCGTTTAAGAGGTTTCTTTGAGAAGGTTCTGAACTTACATTAAAACATTTCTTCTTCGCACAGAATGGTGATGCTGc
AAAATATTGAAAGAAGCAAATTCTCCAAAGAAACACTCCAAGACTTGAAGTAATTTTGTAAAGAAGAAGCGTGTCTACCACTACGACG
F Y N F L R L R G F F D E G S E L T L K H F F F A O N G D A A
-Replicase 1b.
GTAAAGATTTTGACTTTTACCGT TATAATAAGCCTACCATTTTAGATATTGTCAAGCTAGAGTTACATATAAGATAGTCTCCGTATT
H---------------------------H-H-------------------------- 3800
ACAATTTCTAAAACTGAAAATGGCAATAATCGGATGGTAAAATCTATAAACAGTTCGATCCAATGTATATTCTATCAGAGAGCAATAA
V K D F D F Y R Y N K P T C C A R W Y K W S R Y
Replicase 1b
TTGACATTTATGAAGGTGGCTGTATTAAGGCATGTGAAGTTGTTGTAACAAATCTTAATAAGAGTGCTGGTTGGCCATAAATAAGTTTGGT
--H------He-H----------------------------H------------------ 13892
AACTGTAAAT ACTTCCACCGACAAATTCCGTACACTTCAACAACATTGTTTAGAATTATTCTCACGACCAACCGGTAATTTATTCAAACCA
F O Y E G G C K. A C E V V V T N N K S A G W P L N K F G
Replicase 1b
AAAGCTAGTTTGTATTACGAATCTATATCTTATGAAGAACAGGATGCTTTGTTTGCTTTGACAAAGCGTAATGTCCTCCCTACTATGACACA
---------------s-s-s-s-s-s-s-s-s-s-s-s-------------------------------------ee-a-a--------- 398
TTTCGATCAAACATAATGCTTAGATATAGAATACTTCTTGTCCTACGAAACAAACGAAACTGTTTCGCATTACAGGAGGGATGATACTGTGT
K A S L Y Y E S S Y E E O D A L. F A L T K R N v L. P T M T O
Replicase 1b
GCTGAACTAAGTAGTATTAGGGTAAAGAACGTCTAGAACTGTGGTGGGTTTCCTGGTCACAATGACACAAGACAATACC
CGACTTAGAATCATACGATAACACCATTTCTTGCACGATCGACAACCACCACAAAGAGACAACAGGTGTTACGGTGTTCGTTAGG
N K. Y. A S G K E R A R T W G G W S L L S T M T T R O Y
Replicase 1b.
ArcAAAAACATCTTAAATCCATTGTTAATACACGCAATGCCACTGTTGTTATTGGT ACTACCAAATTTTATGGTGGTTGGAATAATATGTTG
---------------------------------------a-arease-------------------as-s-s-s---------- 468
TAGTTTTTGTAGAATTTAGGTAACAATTATGTGCGTTACGGTGACAACAATAACCATGATGGTTTAAAATACCACCAACCTTATTATACAAC
H 0 K H L K S V N T R N A T V V E G T T K F Y G G W N N M
Replicase 1b.
CGT ACTTAATTGAGGFGTTGAAAACCCTATGCTCATGGGTTGGGATTATCCCAAATGTGATAGAGCTTTGCCTAACATGATAC GTATGAT
------------------------------------------------------------------------------------ 1426O
GCAGAAATAACTACCACAATTTGGAACGAG ACCCAACCCTAATAGGGTTACACATCTCGAAACGGATTGTACTATGCATACTA
R T L D G V E N P M L M G W D Y P K C D R A P N m R M
Replicase 1b.
TTCAGCCATGGTGTTGGGTTCTAAGCATGTTAATTGTTGTACTGTAACAGATAGGTTTTATAGGCTTGGTAACGAGTTGGCACAAGTTTTAA
-------------------------a-a-a-a-la-e-eles-o-o-en-le--------------------as-s-s-s-s-s--s-s-s-s-s-s---s-s-s-s-s- 14352
AATCGGT ACCACAACCCAAGA CGTACAATTAACAACATGACATGTCTATCCAAAAATCCGAACCATfGCTCAACCGTGTTCAA AATT
s A M W S K V N C C T W T - D R F Y R G N E A O. W. L.
Replicase 1b.
CAGAAGTTGTTT ArtCAAGGGGTTTTTATTFTAAGCCAGGTGGTACGACTTCGGTGACGCTAGTACAGCTTATGCTAATTCTATTT
------------------------------------------------------------------------------ 1444
GCTCAACAAAAAGATTACACCAAAAAAAAATCGGCCACCATGCTGAAGACCACTGCGATCATGTCGAATACGATAAGATAAAAA
T E V V Y S N G G F Y F K P G G T T S G D A S T A Y A N S I F
Replicase 1b
AACATTTTTCAAGCCGTGAGTTCTAACATTAACAGGTTGCTTAGTGTCCCATCAGATTCATGTAATAATGTTAATGTTAGGGATCTACAACG
------------ee-------------alla-la-----as-------------------------------------- 14536
TTGAAAAATCGGCATCAAGATTGAATGTCCAA CGAATCACAGGGTAGTCTAAGTACATTATACAATTACAATCCCTAGAGTFGC
N F O A v S S N N R S W P S D S C N N W. N. W. R D - O R
Replicase b
ACGTCTGTATGATAATTGCTATAGGTT AACTAGTGTTGAAGAGTCATTCATTGATGATTATTATGGTTATCTTAGGAAACATTTTTCAATGA
--------------------------------------as-s-s-el-res-------------------ee-ee---------------- 4628
TGCAGACATAC Art AACGATATCCAATGATCACAACTTCTCAGTAAGAACTACTAATAATACAAT AGAATCCTTGAAAAAGTACT
R Y D N C Y R L T S W E E S F | D D Y Y G Y L R K H F S M
Replicase 1b.
TGATTCTCTCTGATGACGGTGTTGTCTGTATAACAAGGATTATGCTGAG TAGGTTATATAGCAGACATTAGTGCTTTTAAAGCCACTTTG
-------------------------------all-------------------------------------------------- 1720
AcTAAGAGAGACTACTGcCACAAcAGACAATATTGTTCCTAAT ACGACT CAATCCAATATATCGTCTGTAATCACGAAAATTTCGGTGAAAC
M L S D D G V V C Y N K D Y A E L G Y A D F S A F K A T L.
Replicase 1b.
TATTACCAGAATAATGTCTTTATGAGTACTTCTAAATGTTGGGTTGAAGAAGATTTAACTAAGGGACCACATGAGTTTTGT TccCAGCATAC
------------------------------------------------------ 14812
ATAATGGTCTTATTACAGAAATACT CATGAAGATTTACAACCCAACTTCTTCTAAATTGATTCCCTGGTGTACT CAAAACAAGGGTCGTATG
Y Y O N N V F M S T S K C W W E E D L T K G P H E F C S 0 H, T
Replicase 1b.
TATGCAAATAGTTGATAAAGATGGTACCTATTATTT GCCTTACCCAGATCCTAG TAGGATCTTGTCAGCTGGTGTTTTTGTTGATGATGTTG
H-r------------------------------Ha-------------- 4904
ATACGTTATCAACTATTCTACCATGGATAATAAACGGAATGGGTCTAGGATCATCCAGAACAGTCGACCACAAAAACAAcTACTACAAC
M O V 0 K D G T Y Y P Y P o P S R L S A G V F W D D V
Replicase 1b.
TTAAGACAGATGCTGTTGTTTTGT TAKAACGTTATGTGTCTTAGCTATTGATGCATACCCTCTTTCAAAACACCCTAATTCTGAATATCGT
H-H-----------------------------------H 4996
AATTCTGTCTACGACAACAAAACAATMTTGCAAT ACACAGAAATCGATAACTACGTATGGGAGAAAGTTTTGTGGGATTAAGACTTATAGCA
V K o A V V R Y V S L A D A Y P L S K H P N S E Y R
Replicase 1b
AAGGTTTTTT ACGTATTACTTGATTGGGTTAAGCATCTTAACAAAAATTTGAATGAGGGTGTTCTTGAATCTTTTTCTGT TACACTTCTTGA
-------------------------------------H----H------------------------ 15088
TTCCAAAAAATGCATAATGAACTAACCCAATTCGTAGAATTGTTTTAAACTTACTCCCACAAGAACTTAGAAAAAGACAATGTGAAGAACT
K V F - Y v L L D w w K H L N.Replicase
K. N. 1b. N E G W L E S F S v T L L D
TAATCAAGAAGATAAGTTTTGGTGTGAAGATTTTTATGCTAGTATGTATGAAAATTCTACAATATTGCAAGCTGCTGGCTTATGTGTTGTTT
ATTAGTTCTTCTATTCAAAACCACACTTCTAAAAATACGATCATACATACTTTTAAGATGT TATAACGTTCGACGACCGAATACACAACAAA
N. O. E. D. K. F W C E O F Y A S M Y " E N S T. L. O. A. A. G. L. C. W. W
Replicase 1b
GGGTTCACAAATGTCTTCGTTGGGTGATTGTCTGCGTAAGCCTAGTTGTGCATAAATGGCATATGATCAGTATTGGTACCGAC
---He---a---------------------------------------------------------- 15272
CACCAAGTGTTTGACAAGAAGC AACACCACTAACAGACGCATTCSGATACAACACGTGATTACACGTATACTAGTACATAAACATGGCTG
C G S O T W L R C G D C L R K P C T K C A Y O H. W. F. G T D
Replicase 1b.
CACAAGTTTATTTTGGCTATAACACCGTATGTATGTAATGCATCAGGTTGTGGTGTTAGTGATGTTAAAAAATTGTATCTTGGTGGTTTGAA
GTGTTCAAATAAAACCGATATTGTGGCATACATACATTACGTAGTCCAACACCACAATCACTACAATTTTTAACAAGAACCACCAAACTT
H K F A T P Y W C N A S G C G W S D V K K L Y G G N
Replicase 1b.
TTACTATTGACAAATCATAAACCACAGTTGTCTTTTCCATTATGTTCGCTGGTAATATATTGGTTTATATAAAAATTCAGCAACTGGT
---H-----------He-or-----------------------H 15458
AATGATAACATGTTTAGTATTGGTGTCAACAGAAAAGGTAATACAAGACGACCATTATATAAACCAAATATATTTTTAAGTCGTTGACCAA
Y Y C T N H K P O L S F P - c s A G N F G Y K N S A T G
Replicase 1b.
CCTTAGATGTTGAAGTTTTTAATAGGCTTGCAACGTCTGATTGGACTGATGTTAGGGACTATAAACTTGCTAATGATGTTAAAGATACACTT
GGAATCTACAACTTCAAAAATTATCCGAACGTTGCAGACTAACCGACACAATCCCTGATATTTGAACGATTACTACAATTTCTATGTGAA
S D V E V F N R A S D W T D W R D Y K. L. A. N. D. W. K. D. T. L.
Replicase 1b
AGACTCTTTGCGGCTGAAACTATTAAAGCTAAAGAAGAGAGTGTTAAGTCTTCTTATGCTTTTGCAACTCTTAAAGAGGTTGGGACCTAA
-------------------------------------ree----as-s-s-s-s-s-s-s------------------------------------ 15640
TCTGAGAAACGCCGACTTTGATAATTTCGATTTCTTCTCTCACAATTCAGAAGAATACGAAAACGTTGAGAATTTCTCCAACAAccTGGATT
R L. F A A E T K. A K E E S-Replicase
v K 1b S S Y A F A T L K E v. v G P K
AGAATTGCTTCTTAGTTGGGAAAGTGGTAAAGTTAAACCACCTTTGAATCGTAATTCTGTTTTCACCTGTTTTCAAATAAGTAAGGACT CAA
TCTTAACGAAGAATCAAccCTTTCACCATTTCAATTTGGTGGAAACTTAGCAT TAAGACAAAAGTGGACAAAAGTTTATTCATTCCTGAGTT
E L L L S W E S G K V K P P L N R N S v F T C F O S K D S
Replicase 1b.
AATTCCAAATAGGTGAGTTCATCTTTGAAAAGGTTGAATATGGTTCTGATACTGTTACGTATAAGTCTACTGTAACCACTAAGTTAGTTCCT
---------------------------------------------------------------------------- 15824
TTAAGGTTTATCCACTCAAGTAGAAACTTTTCCAACTTATACCAAGACTATGACAATGCATATTCAGATGACATTGGTGATTCAATCAAGGA
K F O G E F F E K W E Y G S D T W T Y K S T W T T K L V P
Replicase 1b
GGTATGATTTTTGTCTTAACATCTCACAATGTTCAACCTTTACGTGCACCAACTATTGCAAACCAAGAGAAGTATTCTAGCATTTATAAATT
a--------------------------------ee-r-s-s-s-s---------------------H----------H-- 15916
CCATACTAAAAACAGAATTTAGAGTGTTACAAGTGGAAATGCACGGGTTGATAACGTTTGGTTCTCTCATAAGATCGTAAATATAA
G M F W L T S H N V O P L R A P T A N O. E. K. Y S S Y K
Replicase 1b.
GCACCCTGCTTTTAATGTCAGTGATGCATATGCTAATTTGGTTCCATATT ACCAACTTATTGGTAAACAAAAGATAACTACAATACAGGGTC
----------------------------------s-s-s-s-s-s---------------------------------- 6008
CGTGGGACGAAAATTACAGTCACTACGTATACGATTAAACCAAGGTATAATGGTTGAATAACCATTTGTTTTCTATTGATGTTATGTCCCAG
H P A F N W S D A Y A N W P :Y Y O L G. K. 0 K T O G
Replicase 1b.
CTCCTGGTAGTGGTAAGTCACATTGTTCCATTGGACTTGGATTGTACTATCCAGGTGCGCGTATTGTTTTTGTTGCTTGTGCCCATGCTGCT
GAGGACCAT CACCATTCAGTGT AACAAGGTAACCTGAACCTAACATGATAGGTCCACGCGCATAACAAAAACAACGAACACGGGTACGACGA
P P. G. S G K S H C S G L G Y Y P G A R V. F W A A A A
Replicase 1b
TGATTCCTATGGCAAAACAGATGTTTATAGCATTGATAAGTGACTAGGATATACCTGCAAGAGCTCGGGTTGAGGTATAG
---------------------a--as-a-a-a-a-a-i-saa-ee--------ee---------------------------------------ee------ 1692
CAACTAAGGAATACACGTTTTCGATACTGACAAATATCGTAACTATTCACATGATCCTAATATGGACGTTCTCGAGCCCAACT CACAATATC
W D S L c A K A M T W Y S K C T R P A R A R W E C Y S.
Replicase 1b.
TGGCTAAAccAAAAACACAGTGCACAATACATATTTACACTGTAACGCAAccGAGTGTAATGCTGAATGTGTTGAGATs
-------------------ses------------------------reles-s-s-s---------------------------------------- 628.
ACCGAAATTTGGTTTATTGTGATCACGGTTATGTAAAATCGGACAATGcGTAAGGACACATTACGACTAAAcAACAACATCTAC
G F x P N N T S A Y f S T W N A L P E c N A D w w w
Replicase 1b
AAGTTTCAATGTGTACAAATTATGACCTTTCTGTTATTAATCAGCGTTTATCATATAAACATATTGTTTATGTTGGTGATCCACAACAACTT
------------------------------------eeee------------ror---------------------------eler-see----- 6376
CAAAGTACACATGTTTAATACTGGAAAGACAATAATTAGTCGCAAATAGTATATTGTATAACAAATACAACCACTAGGGTTGTTGAA
E v. S M C T N Y O L S v i NO R L 1b
-Replicase S Y K H 1 v Y V G D P o O L
CCTGCACC TAGAGTAATGATTACTAAAGGTGTTATGGAGCCTGTTGATTATAACGTTGTTACTCAACGTATGTGTGCTATAGGCCCTGATGT
GGACGTGGATCTCATTACTAATGATTTCCACAATACCTCGGACAACTAATATTGCAACAATGAGTTGCATACACACGATATCCGGGACTACA
P A P R V M T K G W M E P W D Y N W W T O R M C A G. P. D. W.
Replicase 1b
TTTTCTTCATAAATGTTAT AGATGTCCTGCTGAAATAGTTAATACAGTTTCTGAACTTGTTTATGAGAACAAGTTTGTCCCTGTTAAAccTG
are--------------------------------------------------------------------------------------- 16560
AAAAGAAGTATTTACAAT ATCTACAGGACGACTTTATCAATTATGTCAAAGACTTGAACAAATACT CTTGTTCAAACAGGGACAATTTGGAC
F H K C Y R C P A E I V N T W S E W Y E N K F W P V K P
Replicase 1b.
CTAGTAAACAGTGTTTTAAAATTTTTTTAAGGGTAATGTACAGGTTGACAATGGCTCTAGTATTAACAGAAAGCAGCTTGAAATAGTTAAG
----------------------------------He--------------------- 16652
GATCATTT GTCACAAAATTTTAGAAAAAATTCCCATTACATGTCCAACTGTTACCGAGATCATAATTGTCTTTCGTCGAACTTTATCAATTC
A S K O C F K F. F. K. G. N. V 0 V D N. G. S S N R K 0 W K
Replicase 1b.
CTGTTTTTAGTTAAAAATCCAAGTGGAGTAAGGCTGTGTATTTCTCCTTATAATAGTCAGAATTATGTTGCAGTAGATTTTTAGGACT
--------------------------------H---- 16744
GACAAAAACAATTTTTAGGTTCAACCTCATTCCGACACAAATAAAGAGGAATATTATCAGTCTTAATACAACGATCATCTAAAAATCCTGA
F L V K N P S W S K A V F S P Y N S O N Y V A S R F - G -
- Replicase 1b
TCAAATTCAAACTGTTGATT CTTCT CAAGGTAGTGAGTATGATTATGTAATCTATGCACAAACTTCTGACACTGCACATGCTTGCAATGTAA

-------------------------------H4------------------------------------- 16836
AGTTAAGTTTGACAACTAAGAAGAGTTCCATCACTCATACTAATACATTAGATACGTGTTTGAAG ACTGTGACGTGTACGAACGTACATT
O O T W D S S O G. S E Y D Y V Y A. O. T S D T A H A C N W
Replicase 1b
ACCGTTTTAATGTGCTATAACACGTGCTAAGAAGGGTATATT TTGTGTAATGTGTGATAAAACTTT GTTTGATTCACT TAAGTTTTTTGAG
----------------------------------------one-o-callele------------Hees-le-His-Heels- 16928
TGGCAAAATTACA ACGATATTGGCACGATTTTCCCATATAAAACACATTACACACTATTTTGAAACAAACTAAGTGAATTCAAAAAACTC
N R F N V A T R A K K G F C W M C D K T - F D S L K F. F. E
-- Replicase 1b
ATTAAACATGCAGATTTACACTCTAGCCAGGTTTGTGGCTTGTTTAAAAATTGTACACGCACTCCTCTTAATTTACCACCAACT CATGCACA
H---------------------------------------------------- 17020
TAATTTGACGTCAAATGTGAGATCGGCCAAACACCGAACAAATTTTTAACATGGCGTGAGGAGAATTAAATGGTGGTTGAGTACGTGT
k h A O L H S S 0 W C G L. F K N C T :R T P L N P P T H A H
Replicase 1b.
CACTTTCTTGTCGTTGTCAGATCAGTTTAAGACTACAGGTGATTTAGCTGTTCAAAT AGGTT CAAATAATGTTTGT ACTTATGAACATGTTA
H--------------H--------------------------------- 17 12
GTGAAAGAACAGCAACAGCTAGTCAAATTCTGATGTCCACAAATCGACAAGTTTATCCAAGTTTATTACAAACATGAATACTTGACAAT
T F L S L S D C F K T T G D L A V 0 i G S N N v C T Y E H v
-Replicase 1b
TATCATTTATGGGTTTATTGAATTAGTATTCCGGTAGCATAGTTTGTTTGTACACGTGACTTTGCTATTCGTAATGTGGTGs
H --------------H------------------------em-H--------------------------------- 17204
ATAGTAAAT ACCCAAAATCCAAACTATAATCATAAGGACCATCAGTATCAAACAAAACATGTGCACTGAA ACGATAAGCATTACACGCACCA
s F M G F R F D S P G S H S F C T R D F A R N W R G
Replicase 1b
TGGTTGGGTATGGATGTTGAAAGTGCT CATGTTTGTGGCGATAACATAGGTACTAATGTTCCTTTACAGGTTGGTTTTT CAAATGGTGT TAA

ACCAACCCATACCTACAACTTTCACGAGTACAAACACCGCTATTGTATCCATGATTACAAGGAAATGTCCAACCAAAAAGTTT ACCACAATT
W L G M D W E S A H W C G D N J C T N V P L 0 V G F S N G W N
Replicase 1b - -
TTTTGTTGTGCAAACTGAAGGTTGTGTGTCTACCAATTTTGGTGATGTTATTAAACCTGTTT G T GCAAAATCTCCACCAGGTGAACAATTTA
----------------------H------------H---------------------- 17388
AAAACAACACGTTTGACTCCAACACACAGATGGTTAAAACCACTACAATAATTTGGACAAACACGTTTTAGAGGTGGCCACT GTTAAAT
F W W G T E G C V S T N F G D v K P V C A K S P P G E O F
Replicase 1b
GACAccTGTTCCTTTTTACGTAAAGGACAACCTTGGTTAATTGTTCGTAGACGCATTGTGCAAATGATATCTGATTATTTTCCAATTG
H------------------------------------------------------------- 1748O
CTGTGGAACAAGGAAAAAATGCATTTCCTGTTGGAACCAATTAACAAGCATCTGCGTAACACGTTTACTATAGACTAATAAACAGGTTAAAC
R H L V P F L R K G O P W L I W R R R V O M I S D Y S N
r-Replicase 1b
TCTGACAT ICTTGTCTTTGTTTTGTGGGCAGGTAGTTTGGAATTAACTACAATGCGT TACTTTGTAAAAATAGGGCCAATTAAATATTGTTA
---------------------------------------------------------------- 17572
AGACTGTAAGAACAGAAACAAAACACCCGTCCATCAAACCTTAATTGAGTTACGCAATGAAACATTTTTATCCCGGTTAATTTATAACAAT
S L. V. F W W A G S L E L T T M R Y F W K G P K Y C Y
Replicase 1b
TTGTGGTAATTCTGCCACTTGT TATAATTCAGTTAGTAATGAATATTGTTGTTTTAAACATGCATTGGGTTGTGATTATGTTTACAATCCGT
AACACCATTAAGACGGTGAACAAT ATTAAGTCAATCATTACT TATAACAACAAAATTTGTACGTAACCCAACACTAATACAAATGTTAGGCA
C G N S A T C Y N S W S N E Y C C F K H A L G C D Y W Y N P
Replicase 1b -
ATGCTTTTGATATACAACAGTGGGGTTATGTTGGTTCCTTGAGCCAGAACCACCACACGTTCTGTAACATTCATAGAAACGAGCATGATGCT
--------------------------Hell-e-His-e----------------------------------------- 17756
TACGAAAACTATATGTTGT CACCCCAATACAACCAAGGAACTCGGTCTTGGTGGTGTGCAAGACATTGTAAGTATCTTTGCTCGTACTACGA
Y A F D F O O. W G Y W G S L S O N H H T F C N E H R N E H D A
Replicase 1b
TCTGGTGATGCTGTTATGACACGTTGTTTGGCAGTACATGATTGTTTT GT CAAAAATGTTGATTGGACTGTAACGTACCCCTTTATTGCAAA
He-H-----------H-H--------------- 17848
AGACCACTACGACAATACTGTGCAACAAACCGTCATGTACTAACAAAACAGTTTTTACAACTAACCTGACATTGCATGGGGAAATAACGTTT
S. G D A W M T R C L A W H D C F V K N. W. D. W T W T Y. P. F. A N
Replicase 1b
TGAGAAATTTATCAATGGCTGTGGGCGTAATGCCAGGGACATGTTGTTCGCGCAGCCTTGAAATTGTATAAACCTAGTGTATTCATGATA
H-----------------------He------------H----------- 1790
ACTCTTAAAAGTACCGACACCCGCATTACAGGTCCCGTACAACAAGCGCGTCGGAACTTTAACATATTTGGATCACAATAAG TACTAT
E K F N G C G R N V G H V V R A A K L Y K P S W H D
Replicase 1b
TTGGTAATCC TAAAGGTGTACGTTGTGCTGTACTGATGCCAAATGGTACTGTTATGACAAGCAACCTGTTAATAGTAATGTCAAGTTGTTG
Hm ------------------------------------------------------ 18032
AACCATTAGGATTTCCACATGC AACACGACAATGACTACGGTTTACCATGACAATACTGTTCGTTGGACAATTATCATTACAGTTCAACAAC
G N P K G V R C A v T D A K W Y C Y D K 0 P v N s N v K L L
Replicase 1b
GATTATGATTATGCAACCCATGGTCAACTTGATGGTCTTTGTTTATTCTGGAATTGTAATGTTGATATGTATccAGAATTTTcAarrergrg
18124
CTAATACT AATACGTTGGGTACCAGTTGAACTACCAGAAACAAATAAGACCTTAACATTACAACTATACATAGGTCTTAAAAG TAACACAC
O Y D Y A T H G O L. D G L C. L. F W N C N v D M Y P E F s v c
Replicase 1b - - -
TCGCTTTGACACACGTACTCGTTCTGTTTTTAATTTAGAAGGTGTTAATGGTGGTTCTCTTATGTTAACAAACATGCGTTTCATACAccA
--------------------Hea------------m-e-Hors------------------------------------------ 18216
AGCSAAACTTTGCATGAGCAAGACAAAAAAAATCTTCCAAATTACACCAAGAGAAATACAATTGTTTGTACGCAAATATTGGTC
R F D T R T R S W F N E G V N G G s Y V N K H A F P
Replicase 1b.
CATATGATAAACGTGCTTTTGTTAAATTAAAACCTATGCCCTTTTTTTACTTTGATGACAGTGATTGTGATGTTGTGCAAGAACAAGTTAAT
Hoi- c- I-ha-------H------------------- 18308
GTATACTATTT GCACGAAAACAATTTAATTTTGGATACGGGAAAAAAATGAAACTACTGTCACTAACACTACAACACGTTCTTGTTCAATTA
A Y O K R A F W K K P M P F F Y F D O S D C D W W C E O W N
Replicase b
TATGTACCCCTTCGCGCTAGTAGTTGTGTACCCGTTGTAATATAGGGGTGCTGTTTGTTCAAAACATGCAAATTGTATCAAAAATATGT
H--------------------------- 8400
ATACATGGGGAAGCGCGATCATCAACACAATGGGCAACATTATATCCACCACGACAAACAAGTTTTGTACGTTTAAACATAGTTTTTATACA
Y W P L R A S S C W T R C N G G A V C S K H A N Y O K Y W
Replicase 1b
TGAGGCATATAATACATTTACACAGGCTGGTTTTAACATTTGGGTACCACATAGTTTTGATGTTTATAATTTGTGGCAAATTTTTATTGAAA
ACTCCGTATATATGTAAATGTGTCCGACCAAAATTGAAACCCATGGFGTATCAAAACTACAAATATAAACACCGTTTAAAAATAACTTT
E A Y N T F T O A G F N I w w P H S F D V Y N L W 0 1 F E
Replicase 1b---
CTAATTTACAAAGTCTTGAAAATATAGCATAATGTTGTAAAAAAAGGGTGTTTTACTGGGTTGAGGGAGTTACCTGTTGCAGTTTT
---------------He---H---------H--------- 1858
GATTAAATTTCAAACTTATATCGAAAACAACATTTTTTCCCACAAAATGACCACAACTACCATCAAGGACAACGCAACAA
T N O S E N A F N V V K K G C F T G W D G E L P W A V W
Replicase 1b
AACGACAAAGTTTTTGTTCGCTATGGCGATGTTGACAACTTGGTTTTTACAAATAAAACAACATTGCCTAcTAATGTTGCTTTTGAATGTT
H-------------------------------------H-------- 8676
TTGCTGTTTCAAAAACAAGCGAACCGCTACAACTGTTGAACCAAAAATGTTTATTTTGTTGTAACGGATGATTACAACGAAAACTTAACAA
N D K v F W R Y G D v D N L. v F T N K T T L P T N v A F E F
Replicase 1b.
TGCAAAACGAAAAAGGGTTTAAACAATTGTCTATTCTCAAAAATCTTGGTGTTGTGCACATAAAATTTGTTTTATGGGATTATG
e -H-Ho-H-------------- 18768
ACGTTTTGCTTTTTAccCAAATTGTGGTGGTAACAGATAAGAGTTTTTAGAACCACAACAACGATGTATATTTAAACAAAAT ACCCTAATAC
A K R K M G L T P P L S. L. K. N. L. G. W. W. A T Y K F W L W D Y
Replicase 1b
AAGCTGAAAGACCTTTTAccTCATATACTAAGAGTGTAGTAAATACACTGATTTTAATGAGGATGTTTGTGTTTGTTTTGACAATAGTATT
--------------------------------------------------------------------------------------- 188so
TTCGACTTCTGGAAAATCGAGTATATGATTCTCACATACATTTATGTGACTAAAATTACTCCACAAACACAAACAAAACTGTTATCATAA
E A E R P F T S Y T K S W C K Y T D F N E D W C v c F D N S
Replicase 1b.
CAGGGTTCGTATGAGCGTTTTACGCTTACTACGAACGCTGTTTTATTTTCTACTGTTGTCATTAAAAATTTAACAccTATAAAGTTGAATTT
------------------------------------------------------------- 8952
GTCCCAAGCATACTCGCAAAAYGCGAATGATGCTTGCGACAAAATAAAAGATGACAACAGTAATTTTAAATTGTGGATATTTCAACTTAAA
O G S Y E R F T L T T N A v L. F S T V V K N L T P K L N F
H Replicase 1b
TGGTATGTTGAATGGTATGCCAGTTTCTTCTATTAAGAGTGATAAAGGTGTTGAAAAATTAGTTAATTGGTACACAT ATGTTCGTAAAAATG
--------------------real-les-----as-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-s-trs-s-s-s-s-s------------------- 90
ACCAACAACTTACCATACGGTCAAAGAAGATAATTTCACTATTTCCACAACTTTTAACAATTAACCATGGATACAAGCATTTTAC
G M N G M P W S S K S D K G W E K L V N W Y T Y W R K N
-Replicase 1b.
GTCAATTTCAAGATCATATGAGTTTTACACTCAAGGTAGGAATTTATCAGACTTACACCAAGAATGATATGGAGTATGATTTC
---------------------------------------------------ee--------------------------- 1936
CAGTAAAGTCTAGTAATACTACCAAAAATGTGAGTCCATCCTTAAAAGTCFGAAATGTGGTTCTTCACA ACCCATACAAAAGAA
C F O D H Y D F Y T O G R N S D F T P R S D M E Y D F
--Replicase 1b
AACATGGATAGGGGTTTATTAATAAAAGGTCTTGAGGATTAATTGAACAGTTGTATATGGTATTTCAAAAACTACA
-----------------------Hell------------------------------- g228
TTGTACCTATACCCACAAAAATAATTATTTATACCAGAACTCCTAAAATTAAAACTTGTACAACATATACCACTACAAAGTTTTTGATGTAA
N M D M G V F N K Y G L E D F N F E H W W Y W S K T L
Replicase 1b
AGGAGGT CTTCATTTGTTGATATCACAGTTTAGGCTTAGTAAAATGGGTGTTTTGAAAGC TGATGATTTTGTCACTGCTTCTGACACAACTT
--------------------He---------Hi-------------------- 932O
TCCCCAGAAGAAAAACTATAGTCAAACCGAATCATTTTACCCACAAAACTTCGACTACAAAAAGGACGAAGATTTTGAA
G G S O F R S K M G W L .K A D D F W A S D T
-Replicase 1b.
TGAGGGCTGTACTGF ACTTACTTAATGAACTAGTTcAAAAGGTTTGTACAATGGATTTGGTTGACGACTTTTTACTATA
ACTCCACGACATGACAATGAATAGAATTACTTGAATCAAGTTTTCAACAAACATGAATATACCTAAACAACAACCTGCTGAAACAATGATAT
L R C C T W T Y. L N E L S S K V V C T Y. M. D. L. L. L. D. D F W T
-Replicase 1b.
CTAAAGAGTTTAGATCTTGGTGTAATATCTAAAGTTCATGAAGTTATTATAGATAATAAACCTTATAGGTGGATGTTGTGGTGTAAAGATAA
re-r--Hire------------------------------------------- g50
GATTTCTCAAATCTAGAACCACAAT AGATTCAAGTACTTcAAAAACTATTATTGAAATCCACCTACAACACCACATTTCTATT
K S L O L G V S K W H E V D N K P Y R W M L W C K O N
-Replicase 1b.
CCACTTGTCATTACCACAGTTGAGCTGCTGAATGGAAGTGGGTTATGCTAGCCACAAATTATAACTTCAACGTATTGT
GTGAACAGCTGAAAAAAGGTTCAACGCAGACGACTTACCTTCACACAATACGATACGGTTTTAAATATCAAGTCATACACAA
H L S T F Y P O L O S A E W K C G Y A M P O K O R M C
- H --Replicase 1b.
GAACTTGTAATATAAATTATCGGTATAATTCCTATAAATTTAAATGGTTAAAACACCAGCTTGTCAA
-------------------------------------------------------------------------------------------- 19688
AccGSAACATAAATATATTAATACCACGACCATAATTCAACGGAT CACCATATTACAATTTACAACAATTATGTGAGTCGAAACAGTT
L. E P C N Y N Y G A G K P S M L N V V K Y T C
Replicase 1b
TACCTAAATAGCACACAAGTSCGTACCTCATAAATGCGGTTTTGATATGGTGCGGTCTGACAAAGGTTGGCACCTGGTACAAC
----------------------------------------------ar-e------------------------------------ g780
ATGGATTTATCGTGATGTTACACGCATGGAATTATACGCACAAAACGTGATACACACCAAGACTGTCCACACCGTGACCATTT
Y L. N S T C. V P H. N. M. R W L H Y G A G S D K G w A P G T T
m Replicase 1b.
TGTTTTAAAACGTTGGCTACCACCTGATGCAATAATCATGATAATGAACAATGATATGTAGTATGCAGATTTTAGCATTACAGGTG
s-----------------a-a----ee-cre--series------------------------------es------------------as 1972
ACAAAATTTTGCAACCGAGGGGACTACGTTATTAGAACTATTACTAAGTTACTAATACAACACTACGCTAAAATCGTAAGTCCAC
v L. K R W L P P D A D N D N D Y V S D A D F S T G
Replicase 1b.
ATTGTGCTACTGTTTAccTTGAAGATAAGTTTGACTTACTTATTTCTGATATGTATGATGGTAGAATTAAATTTTGTGATGGTGAAAACGTC
----------------------------------Helles-s-s-s- 19964
TAACACGATGACAAAGGAACTTATTCAAACAATAATAAAACTATACAAcTACCATCTTAATTTAAAAACTACCACTTTGCAG
A T W Y K F D L S o M. Y D G R K F C D G E N W
Replicase ib
TCTAAAATGGTTTTTTACTACTTAATGGTGTATTAGAAAAAATTAs. ATGGGGTATGTTCCATTAAGATTACAGAATATAG
---------------------------------------------------------------------He----------- 20056
AGATTACCAAAAAAATGAAT AGAATTACCACAATAATCTCTTTTTAATCGATAACCACCATCACAACGGTAATTCTAATGCTTATATC
s K D G F F T Y. L N G v I R E K L A G G S v A K T E Y S
-Replicase 1b.
TGGAAAAGTACTTAGAATAATACAAAGATTTGCTTTTTGGACTTTGTTCTGCACGTCTTTAATACACCTCTCAGAAGCTTTC
-------------------------------as-s-s-s-s-as-s-s-s-s-s-s-asser----ee-le-ses----------------- 20148
AACCTTATTCATAGAAAT ACTTAATTATGTTTCTAAACGAAAAACCTGAAACAAGACGTGCAGACAATTATGTAGGAGAAGTCTTCGAAAAG
W N. K. Y Y 0 R F A F W T L F T S V N T S S S E A F
Replicase 1b
TTAftgGTATTAATTATTTAGGTGACTTTATTCAAGGTCCTTTTATAGCTGGTAACACTGTTCATGCTAATTATATATTTTGGCGTAATTCT
---------------------------all-all-Hass-s-s-s-s-s-s-s-s-s-s-s-rele-------------------------ee-no-ree- 2:240
AATAACCATAATTAAAAATCATAAATAATTCCAGGAAAATATCGACCATTGTGACAAGACGATTAATATATAAAACCCATTAAGA
L N Y O G. P. F. A G N T W H A N Y F. W R N S
Replicase 1b
ACTATTAGTCTTGTCATACAATTCAGTTTTAGATTTAAGAAGTTTGAAGTAAACATA AGGCCActg. GTGTTACATAAAGATAG
TGATAAT ACAGAAACAGTATGT TAAGTCAAAATCTAAATTCATTCAAACTTACATTTGTATTCCGGTGACAACAACAATGTGAATTTCTATC
T M S L S Y N S v L. D. L. S.Repticase
K. F. 1b.E. C K H K A T V V V T L K D S
TGATGTAAATGATATGGTTTGAGTTfGATTAAGAGTGGTAGGTTGTTGT Act AATAGTGGCCGTTTTGGTGGTTTAGTAATCATTTAG
al-e-------------------------------------------------------------------------r- 2024
ACACATTACAt ACCAAAACCAAACAATTCTCACCATCCAACAACAATGCAFTATCACCGGCAAAACCACCAAAATCATAGTAAATC
O W N D M W L S K s G R l l R N S G R F G G F S N
-Replicase 1b
TCT CAACTAAATGAAACTTTTCTTGATTTTGCTTATTTTGCCCCTGGTTTCTTGCTTTTCTACATGTAACAGTAATGCTAGTATT ICTATGT

20516
AGAGTTGATTT ACTTTGAAAAGAACTAAAACGAATAAAACGGGGACCAAAGAACGAAAAGATGTACATT GTCATTACGATCATAAAGATACA
M K L F L I L L T L P L v S C F S T C N S. N. A s 1 s M
- spike - - - - - - - - - - - -
W S T K .
-Replicase 1b.
TACAATTAGGTGTTCCTGATAACTCTTCAACTATTGTCACAGGTTTGTTGCCAGTCCATTGGATTT GTGCTAATCAGAGTACATCTAGTTAC
-------------------------------- 20608
ATGTTAATCCACAAGGACTATTGAGAAGTTGATAACAGTGTCCAAACAACGGTCAGGTAACCTAAACACGATTAGTCTCATGTAGATCAATG
L O L G V P D N S S T W T G L L P V H W I C A N Q S T S S Y
Spike
CCAGCCAACGGCTTTTTCTATATTGAGTGGTAAACACCGTAGTGCCTTGCACTCCATAGTGGTATTATGATGCTAACCAGTATTATA
----------------H----------------Henri-----------rmer-He-Hen-Ho-Haar 20700
GGTCGGTTGCCGAAAAAGATATAACTACAACCATTTGTGGCATCACGGAAACGTGAGGTAT CACCAATAATACTACGATTGGTCATAATATA
P A N G F F Y D. W. G K H R S A F A L H S G Y Y O A N 0 Y Y
-Spike
TTACT CACTAATAAAATACATTAAATGCTCCTGTCACTCTGAAGATTTGTAAGTTTGGAAACACTTCTTTTGATTTTTTAAGTAATGTTT
-------------------------------------------------------------------------------- 20792
AAAGAGTGATTATTTTATGTAAATTTACGAGGACAGTGAGACTTCTAAACATTCAAACCTTGTGAAGAAAACTAAAAAATTCATTACAAA
Y T N K H N A P W T L K C K F. G. N T S F D F L S N V
Spike
CTACTTCTCATGATTGTATAGTTAATTT GTCATTCACAGAACAGTTAGGTGTGCCTTTGGGCATAACTATATCGGGTGAAACTGTACGTTTG
H---------------------------------H------- 20884
GATGAAGAGTACTAACATATCAATTAAACAGTAAGTGT CTTGTCAATCCACACGGAAACCCGTATTGATATAGCCCACTTTGACATGCAAAC
S T S H D C V N S F T E O L G W P L G T J S G T W R L.
Spike
CATTTATATAATGCAACTCGTACTTTTTATGTGCCGGCCGCTTATAAACTTACTAAACTTAGTGTTAAATGTTAcTTTAGTGAATCCTGTGT
H----------------------------------------------------------- 20976
GTAAATATATTACGTTGAGCATGAAAAATACACGGCCGGCGAATATTTGAATGATTTGAATCACAATTACAATGAAATCACTTAGACACA
H L Y N A T R T F Y V P A A Y K L T K L S W K C Y F S E S C V
Spike
TTTTAGTGTTGTCAATGCCACCATTACTGTTAATGTCACCACACTTAATGGCCGTATAGTTAACTACACTGTTTGTGATGATTGTAATGGTT
-----------------------H---------------------------------------------------------- 2068
AAAATCACAAATTACGGTGGTAATGACAATTACAGTGGTGTGAATACCGGCATACAATTGATGTGACAAACACTACTAACATTACCAA
F S W W N A T T V N W T T N G R W. N. Y. T W C D D C N G
Spike
ATACTGATAACATATTTTCTGTTCAACAGGATGGCCGCATTCCTAATGGTTTCCCTTTTAATAATTGGTTTTTGTTAACTAATGGTTCCACA
---------------all-a--------------------------------------------------- 21 160
ATGACATTGTATAAAAGACAAGTTGTCCTACCGGCGTAAGGATTACCAAAGGGAAAATTATTAACCAAAAACAATTGATTACCAAGGGT
Y T D N F S W o o G R P N G F P F N N W F L L T N G s T
Spike
GGTAACCCTCGTTGTAAGAATCTACTTAAGCAGTATACTTCTGCTTGEAAAACTATTGAAGATGCCTTACGACTTAGTGcTCATTTGGAAAC
------------------------------------------------------------------- 22908
CCATTGGGAGC AACATTCTTAGATGAATTCGTCATATGAAGACGAACATTTTGATAACTTCTACGGAATGCTGAATCACGAGTAAACCTTTG
G N P R C K N K o Y T S A C K T . E D A R L S A T
Spike
TAATGAGTTAGAGTATGAACTTCGATAGCAATGCTTTTAGTTTGGCTAATGTTACTAGTTTGGAGA TATAACCTTCTAGTGTT
ATTACTACAATCATCATACGATTGAAAGCTATCGTTACGAAAATCAAACCGATTACAATGATCAAAACCTCTAATATTGGAAAGATCACAAA
N D V S S M L T F S N A F S A N W T S F. G D Y N L S S W
- Spike--
TACT CAGAAAACATTCATTCAAGCCGTATAGCAGGACGTAGTGCTTTGGAAGATTTGTGTTAGAAAGTTTTACATCGGTTTGGG
-----------------------------ee--------as-areasesseries-------------------------------e-r-e-r- 23092
ATGGAGTCTCTTTGTAAGTAAGTTCGGCAT ATCGTCCTGCATCACGAAACCTTCTAAACAACAAATCGTTTCAACAATGTAGACCAAACCCA
P o R N H S S R A G R S A E D L - F S K W W T S G L
- Spike
ACTTGATGTTGACTATAAccTGTACTAAAGGTCTTCATTCTGACCTTGCTTGGCT CAGTACTACAATGGCATAATGGTTTGCC
------------------------------------------------------------------------------ 23 184
TGACAACTACAAcGATATCAGAACATGATTTCCAGAAAGATAACGACGGA ACGAACACGAGTCATGATGTTACCGTATTACCAAAACGG
T v D v D Y K S C T K G L S A D L A C A O Y Y N G M W L P
-Spike
AGGTGTTGCTGATGCTGAACGTATGGCCATGT ACACAGGTTCTCTTATAGGTGGCATGGTGCTCGGAGGTCTTACATCAGCAGCCGCCATAC
-------------------------------------------------------------------------------------- 23276
TCCACAAcGACTACGACTTGCATACCGGTACATGTGTCCAAGAGAATATCCACCGTACCACGAGCCTCCAGAATGTAGTCGTCGGCGGTATG
G v A D A E R M A M Y T G S spike---
J G G M W L G G L T S A A A 1
CTTTTTCTTGCACTGCAAGCACGACTAACTATGGTTTACAAACTGATGTGCTTCAAGAAAATCAGAAAATTGGCTGCATCATTT
GAAAAAGAAAccGTGACGTTCGTGCTGAATTGATACAACGAAATGTTTGACTACACGAAGTTCTTTTAGTCTTTTAAAACCGACGTAGTAAA
P F S A. L. O. A R N Y W A T D V O E N 0 K A A S F
Spike
AATAAGGCATTAATAATATTGTTCTTCTTAGTAGGTTAATGATGTATTACACATACGCAGAGGCTATACATACTGTACTATTGC
-----------------------------------ee-ereo-a-------------------- 2346d
TTATCCGATAATTATATAAAACAAGAAAATCATCGCAATTACACGATAATGTGTAGACTCTCCGAATGATGACAATGATAACG
N. K. A N N W A S F S S V N D A T H T A E A H T W T A
Spike
ACTTAATAAGATTAGGATGTTTTAATAACAGGGTAGTGCTCTAACCATCTCACTTCACAATTGAGACAAATTTTCAGGCCATTCTA
------------------------------ee-----------------------------------------------H------- 23552
TGAAATCTAAGTCCTACAACAAT AGTTGTCCCATCACGAGAATTGAGAGTGAAGTGTTAACCTSTATAAAAGTCCSGTAAAGAT
N K I C D V V N G. S A. N L T S O L R H N F O A S
Spike
ATTCAATTCATGCTArt TAGACCGGCTTGATTCAATTCAAGcGATCAACAATTGACAGATAATTACTGGACGGCTTGCAGCTTGAAT
e------------ee-ee-aa-ee----s-s-s-s-s-s-s-s-s-s-s-s-s-s-reer---as-s-s-s-s-s-s-s---are-his-or-r-er--------------ee---- 236
TAAGTTAAGTACGATAAATACTGGccGAACTAAGTTAAGTTCGGCTAGTTGTTCAAcf GTCTAATTAATGACCTGCCGAACGTCGAAACTTA
N S I H A Y R L D S A o O W D R T G R A A N
-Spike
Fig. 2A
PRCOVTGEV
0.1 EMCR PEDV
Fig. 2B
PED
(31)qg?oaqioploeKxiu?p?iueaenpvaui
|
000" |
000"
ºg-eop&xuaqt(3?luinvo1eapj4enu?ao
SYIVS 000"|
Patent Application Publication Feb. 21, 2008 Sheet 38 of 87 US 2008/004442.6 A1
TCAGGAGGGTGTTTTGTCAGAAAGAGAGAAGTTGCTTCATTTAATCTAAACTAAACAAAATCGCTAGTGTAAATTGGGCCGATGACAGAGCT 26128
AGTCCTCCCACAAAACAGTCTTTCTCTCTTCAACGAAGTAAATTAGATTTGATTTGTTTTACCGATCACAT TIAACCCGGCTACTGTCTCGA
- C - E G - W- L S- E- M--
m E K - L H - . - . -. M. A. S. V--N-
N W A - D D- R - A -
GCTAGGAAGAAATTTCCTCCTCCTTCATTTTACATGCCTCTTTTGGTTAGTTCTGATAAGGCACCATATAGGGTCATTCCCAGGAATCTTGT
---------------------------------------------------------------
CGATCCTTCTTTAAAGGAGGAGGAAGTAAAATGTACGGAGAAAACCAATCAAG ACTATTCCGTGGTATATCCCAGTAAGGGTCCTTAGAACA 26220
A R K K F P P P S F Y M P L N S S D K A P Y R v 1 P R N L v
CCCTATTGGTAAGGGTAATAAAGATGAGCAGATTGGTTATTGGAATGTTCAAGAGCGTTGGCGTATGCGCAGGGGGCAACGTGTTGATTTGC
H------------------------------------------------------------------------ 2632
GGGATAACCATTCCCATTATTTCACTCGTCTAACCAATAACCTTACAAGTCTCGCAACCGCATACGCGTCCCCCGTTGCACAACTAAACG
P e K. G. N. K D E O G Y W NY O E R W R M R R G O R W D
CTCCAAATCATTTATTACCAGGTACTGGACCTCAAAGGACCTTAAATTCAGACAAcGTTCTGATGGTGTTGTTTGGGTTGcTAAG
GAGGATTTCAAGTAAAAATAATGGACCATGACCTGGAGTATFCCTGGAATTTAAGTCTGTTGCAAG ACT ACCACAACAAACCCAACGATTC
P P K v H F Y Y L G T G P H K R K F. R O R S D G V V W W A K.
GAAGGTGCTAAAACTGTTAAT ACCAGTCTTGGAATCGCAAACGTAATAAAACCTTTGGAACCAAAGTTCTCTATTGCTTGCCTCCAGA
------------------------------------- 26496
CTTCACGATTTTGACAATTATGGTCAGAACCATTAGCGTTTGCATTAGTCTTTGAAACCTTGGTTTCAAGAGATAACGAAACGGAGGTCT
E G A K T V N T S L G. N. R. K. R O K P L E . P K F S A P P E
GCTCTCTGTTGTfGAGTTTGAGGATCGCTCTAATAACTCATCCGTGCTAGCAGTCGTTCTTCAACTCGTAACAACTCACGAGACTCTCTC
her---------------------H-----------------H--------------------------
CGAGAGACAACAACT CAAACTCCTAGCGAGATTATTGAG TAGAGCACGATCGTCAGCAAGAAGTTGAGCATTGTTGAGTGCTCTGAGAAGAG 26588
S W W E F E D R S N N S S n S S R S S T R N N S R D S S
GTAGTACT TCAAGACA ACAGTCTCGCACTCGTTCTGATTCTAACCAGTCTTCTTCAGAT CTTGTTGCTGCTGTTACTTTGGCTTTAAAGAAC

CATCATGAACTTCTGTTGTCAGAGCGTGACAAGACTAAGATTGGTCAGAAGAAGTCTAGAACAACGACGACAATGAAACCGAAATTCTTG
R S T S R O O S R T R S D S N S S S D L W A A W T L A. L. K. N.
TTAGGTTTTGATA ACCAGTCGAAGT CACCAGTTCTTCTGGTACTCCACCCTAAGAAACCAATAAGCCTCTCTCAACCCAGGGCTGA

-------------H-------------------------------------H-H----------- 26772
AATCCAAAACTATTGGTCAGCTTCAGTGGATCAAGAAGACCATGAAGGTGAGGATTCTTTGGATTATTCGGAGAAAGAGTTGGGTCCCGACT
G F O N O S K S P S S S G T N T P K K P N K P L S o P. R. A. D.
TAAGCCTTCTCAGTTGAAGAAAccTCGTTGGAAGCGTGTTCCTACCAGAGAGGAAAATGTTATTCAGTGCTTTGGTCCTCGTGATTTTAATC
-------Hero-H-------------Hero-Ho-c-e-H-------------Hero-o-o-o-o-H-------------H- 2686
ATCGGAAGAGTCAACTTCTTGGAGC AACCTTCGCACAAGGATGGTCTCTCCTTTTACAATAAGTCACGAAACCAGGAGCACTAAAATTAG
K P S O L K K P R W K R V P T R e N V o c F. G. P R D F N
ACAATATGGGGGATTCAGATCTTGTICAGAATGGTGTTGATGCCAAGGGTTTTCCACAGCTTGCTGAATTGATTCCTAATCAGGCTGcGTTA 26956
TGTTATACCCCCTAAGTCT AGAACAAGTCTTACCACAACTACGGTTCCCAAAAGGTGTCGAACGACTTAACTAAGGATTAGTCCGACGCAAT
H N M G D S D L V O N G v D A K G F P 0 - A E L J P N C A A
TTCTTTGATAGTGAGGTTAGCACTGATGAAGTGGGTGATAATGTTCAGATTACCTACACCTACAAAATGCTTGTAGCTAAGGATAATAAGAA
H----------------H-H--------Hit-------------------- 27048
AAGAAACTATCACTCCAATCGTGACT ACTTCACCCACTATTACAAGTCTAATGGATGTGGATGTTTTACGAACATCGATTCCTATTATTCTT
F F D S E W S T D E W G O N V T Y T Y K M L V A K D. N. K. N
CCTCCAAGTTCATTGAGCAGATTAGTGCTTTACTAAACCCAGTTCTATAAAGAAATGCAGTCACAATCATCTCATGTTCTCAGAACA
------------------------------------------------------------------------------------- 27 140
GGAAGGATTCAAGAACTCGTCTAATCACGAAAATGATTTGGGT CAAGATAGTTTCTTTACGTCAGTGTTAG TAGAG TACAACGAGTCTTGT
P K F E O - S A F T K P in S K E M O S C S S H. W. A N
CAGTACTTAATGCTTCTATTCCAGAATCTAAACCATTGGCTGATGATGATTCAGCCAT TATAGAAATTGTCAACGAGGTTTTGCATTAAATT
---------------------------------------------------------------------------------- 27232
GTCATGAATTACGAAGATAAGGTCTTAGATTTGGTAACCGACTACT ACTAAGTCGGTAATATCTTTAACAGTTGCTCCAAAACGTAATTTAA
. .N . . . . . . . . . . . . . ."
GTTTTGTAATTCCAGTTGAATGTTTATTATTATTAGTTGCAAccCCATGCGTTTAGCGCATGATAAGGGTTTAGTCTTACACACAATGGTAG
------------------------------H-i-H-------------------- 27324
CAAAACAFTAAGGTCAACTTACAAAAATAATAATCAACGTTGGGGTACGCAAATCGCGACTATTCCCAAATCAGAATGTGGTTACCATC
GCCAGTGATAGTAAAGTGTAAGTAATTTGCTATCATATTAACATGTCT AGAGGAAAGTCAGAACTTTTTCTGTTTGTGTTGTTGGAGTACTT
her---------------------------------------------------- 2746
CGGTCACTATCATTCACATTCATTAAACGATAGATAATTGTACAGATCTCCTTTCAGTCTTGAAAAAGACAAACACAACAACCTCATGAA
AAAGATCGCATAGGCGCGCCAACAATGGAAGAGCCAACAACATATCAAAAATGTTTTGTCTGGTACTTGTTAATGATATTGTTTTGATAT
-H---------------------------------H-------H---------------------------- 27508
TTTCTAGCGTATCCGCGCGGTTGTTACCTTCTCGGTTGTTGTATAGATTTTTACAAAACAGACCATGAACAATTACTATAACAAAAACTATA
3'UTR muuum
GGATACACAAAAAAAAAAAAAAAA
-Hees-Hero---> 27.532
CCTATGTGTTTTTTTTTTTTTTTT
mu3'UTRuu
Fig. 4 Alignments
a. 5' untranslated region (Genomic sequence) aligned with human
coronavirus 229E
. ..... . .... ..... . .. ..... . .... ...... . . . .... .... ..... .
5 5 25 35 45 55
EMCR5 UTR - - - - - - - - - - - - - - - - - - - - - - - AGATAGA GAATTTTCTT ATTTAGACT TGTGTCTACT
229E5 UTR ACTTAAGTAC CTTATCTATC TACAGATAGA AAAGTTGCT -TTTAGACT TGTGTCTACT
.. ..... . ........ . .. ..... . .... ...... .. ..... .. ..... .

65 75 85 95 105 15
EMCR5' UTR CCTCTCAACT AAACGAAATT TTT-CTAGTG CTGTCATTTG TTATG--GCA GCCTAGTGT
229E5' UTR TTTCTCAACT AAACGAAAT TTTGCTATGG CCGGCATCTT TGATGCTGGA GTCGTAGTGT
... ..... . .... .. ... .... . .. ..... . .. ..... .. ..... .

25 135 45 55 65 75
EMCR5 UTR AATTGAAATT TCGTCAAGTT TGTAA-ACTG GTTAGGCAAG TGTTGTATTT CTGTGTTTA
229E5 UTR AATTGAAATT CATTTGGGT TGCAACAGT TGGAAGCAAG FGCTGTGTGT CCTA-GTCTA
... ..... . .... ..... . ... ...... .. ..... .. ......

185 95 2O5 25 225 235
EMCR5' UTR AGCACTGGTG GTTCTGTC-C ACTAGTGCAC AC-ATTGATA CTTAAGT-GG TGTTCTGTCA
229E5' UTR AGGGTTTCGT GTTCCGTCAC GAGATTCCAT TCACAAACG CCTTACTCGA GGTTCCGTCT
... ..... . .... ..... .. .. . .. .. . .. .. ... .. . ... .. ....

245 255 265 275 285
EMCR5 UTR CTGCTTATTG TGGAAGCAAC GTTCTGTCGT TGTGGAAACC AATAACTGCT AACC
229E5 UTR CGTGTTTGTG TGGAAGCAAA GTTCTGTCTT TGTGGAAACC AGTAACTTT CCTA
b. Putative Orf a
.... ..... . .. ..... . ..... . .. .. . .... . . . .. ..... .
5 5 25 35 45 55
EMCR - - - - - - - - - - - - -MFYNQVT AWASDSES GFGFAPSWA VRAYSEAAAO GFOACREVAF
229E - - - - - - - - - - - - - MACNRWT AWASDSES ANGCSTIAOA WRRYSEAASN GFRACRFWSL
PEOW - - - - - - - - - - - - - MASNVT AFANDAES AFGFCTASEA WSYYSEAAAS GFMOCREWSL
TGEW --- ---- - - - - - -MSSKOFK ILVNEDYOVN WPSPIRsdw LOEIKYCYRN GFEGYWEVPE
OC43 MSKINKYGLE LHWAPEFPWM EEDAEEKLON PSSSEWOMC STTAOKLETD GICPENHWMW
BoCoW MSKINKYGLE LHWAPEFPWM FEDAEEKIDN PSSSEWDWC STTAOKLETG GICPENHVMW
MHV MAKMGKYGLG FKWAPEFPWM LPNASEKGS PERSEEDGFC PSAAOEPKTK GKTLNHVRW
APW - - - - - - - - - - - - - MASSIKQ GWSPKPROVI LWSKDIPEO COALEETSH NPKDYAOAFA
SARS COW - - - - - - - - - - - - -MESVLG VNEKTHVOLS LPVLQVRDVL WRGFGDSWEE ASEAREHK
........ . .. . .... . .. ..... . .. .. .. . .. ..... .. . .... .
65 75 85 95 05 15
EMCR GLQDCVTGIN DDD-YVIALT GTNCCAKI FSDRPNLR GWLESNSNY VLQDFDVVFG
229E DODCVGA DT-YWMGLH GNOTLFCNIM KFSDRPEMLH GWLVESNSNY LLEEEDWWFG
PEDV DADTWEGILL PEDaYWMWV GTTKLSAYWid TFGSRPRNC GWLFSNCNY FLEELELTFG
GEW YCRDLWDCOR KOH-YWGV, GNGWSDLKPW LLTEPSWMLO GFIVRANCNG WLEDFDLKA
OC43 DCRRLLKOEC CVOSSLIREI WMNASPYDE WLODALOSR EAWLWTTPLG MSLEACYWRG
BoCoW DCRRLLKQEC CWOSSLIREI WMNTRPYDE WLLODALOSR EAWLWTPPLG MSEACYWRG
MHW DCSRLPALEC CWOSAIRDI EWOEPLNWE ASTMMALQFG SAWLWKPSKR LSOAWAKLG
APW VROKFDRSLO TGKOFKFETV CGLELLKGV) KTPG- - - - - WPAKWLKATS KADLEDFG
SARS COW NGTCGLVELE KGVLPOLEOP YWFKRSDAL STNHGHKWWE LVAEMOGOY GRSGTGW.
......... . .... ..... . .. ..... . . . .. .. . . . .. . ... . .

.. ..... .
125 35 145 155 175 1.65
EMCR --HGAGSWWF WDKYMCGFDG KPWLPKNMWE FROYENDNTD S-IWGGVTY OLAWDVIRKD
229E K-RGGGNVTY TDOYLCGADG KPVMSEDLWO FWDHFGENEE --NGHY WCAWTKRKP
PEDW --RRGGNIVP VDOYMCGADG KPVLOESEWE YTDFEADSED GQLNIAGITY WKAWWERSD
TGEW --RTGRGAIY VDQYMCGADG KPVEGs - -D FKDYEGDED aIEEEGEEY HCAWTVRDE
OC43 C-NPKGWTMG FRRRSWCNT GRCTVNKHWA YOLYMIDPAG - - -WCLGAGO FWGWWIPAF
BoCoW C-NPNGWTMG LERRRSWCNT GRCAVNKHWA YOLYMIDPAG - - -WCFGAGO EWGWWPLAF
MHV V-LPKTPAMG FKRFCLCNT RECWCDAHWA FOLFTVQPDG - - -WCLGNGR FIGWEWPWTA
APW WSPLARKYRE LLKTACQWSL TWEALDVRAQ TDEFDPT EILWLOVAAK
SARS COW WPHWGETPA YRNWLLRKNG NKGAGGHSYG IDKSYDLGD EGTOPIEDY EQNWNTKHGS
.... .... . .. . . .... . .... .... . .... .... . .... . . . . . . ... .... .
85 195 2O5 215 225 23S
EMCR SALPPELSV WEFEDRSNNS SRASSRSSTR - - - - - - - - - - - - - - - - NNSR DS- - - - - - - -
229E NQKLPNGVTV WEEPD----- SRAPSRSQSR - - - - - - - - - - ------ SCSR - - - - - - - - - -
PEDW SQQLPSVVEI WEPNTPP--A SRANSRSRSR GNGNNRSRSP SNNRGNNQSR GNSQNRGNNO
TGEW DGKVPGEFOL EVNOS----- -RDNSRSRSO -- - - - - - - - - - - - - - - - - - - - - - - - - - - - -
FeCoW DGKIPPOFOL EVNRS----- -RNNSRSGSQ ------ - - - - - - - - - - - - - - - - - - - - - - - -
PRCOW DGKVPGEFOL EVNOS----- -RDNSRSRSO - - - - - - - - - - - - - - - - - - - - - - - - - - ----
CaCOW DWKWPSEFHL EWNOL- - - - - -RDNSRSRSO ---------- ---------- ----------
RSDACOW APGTVLPGF YVEGS----- GRSAPASRSG - -- - - - - -- - - - -- - - - - - - - - - - - --- --
MHW APGTVLPQGF YWEGS----- GRSAPASRSG - - - - - - - -- - -- - - - - - - - - -- - - - - - - --
PHEW PPGTVLPOGY YIEGS----- GRSAPNSRST - - - - - - -- - - -- - - - - - - - - -- - - - - - - - -
OC43 PPGTVLPOGY YIEGS ----- GRSAPNSRST - - - - - a - a-- --- - - - - - - - - - -- - --- - -
BoCoW PPGTWLOGY YIEGS----- GRSAPNSRST --- - - - - - - - -- - - - - - - - - --- - - - --- -
SARS PQGTTLPKGF YAEGS----- -RGGSOASSR -- -- -- -- -- -- - - - - - - - - -- -- - - ----
ABW SDGGPDGNER WDEP- - - - - - - LN-RGRSG -- - - - - - - - - -- - - - - - - - - - - - - - - --- -
.. ..... . .. .. .... . .... ... . . .... . .. .. . .. . . . .... . . . .. . . ... .

245 255 265 275 28S 295
EMCR SRSTSRO-- - - - - - - -OSR- TRSDSNOS- - - - - --S-SDL, VAAVTLALKN LGFDN--QSK
229E GRGESKP- - - - - - - - - OSRN PSSDRNHN- - - - - --SQDD MKAVAAALKS LGFDKP-QEK
PEDW GRGASQNRGG NNNNNNKSRN OSNNRNOSND RGGWTSRDDL, VAAWKDALKS LGIGEN-PDR
TGEW SRSRSRNR- a s- - - as SOSRG ROOFNNKK-- ------ DDSV EQAVLAALKK LGVDTE-KOO
FeCoW SRSWSRNRs - - - - - - SQSRG RHHSNNQ-- - ------NNNV EDTIVAVLEK LGV-TD-KQ
PRCW SRSRSRNR- - - - - - - SQSRG RQQSNNKK-- ------ DSV EOAWLAALKK ILGWYTE-KOO
CaCoW SRSQSRNR- - - - - - - SOSRG RQLSNNKK-- ------ DDNW EOAVIAALKK ILGWDTE-KOO
RSDACOW SRSQSRGP- - - - - - - - NNRA RSSSNORQ- - ----- PASTW KPDMAEEIAA VLAN---LG
MHW SRSOSRGP- - - - - - - - NNRA RSSSNORQ- - - - - - - PASAv KPDMAEEIAA LVLAK---LG
PHEV SRAPNRAPS- - - - - - AGSRS RANSGNRT- - - - - - - SPGW TPDMADOIAS WAK---LG
CC43 SRSSRASS - - - a -ss- AGSRS RANSGNRTse- as - - - - PTSGV TPDMADQIAS LVLAK---LG
BoCoW SRASSRASS or - a sew - AGSRS RANSGNRT-- a- - - - - PTSGV TPDMADOIAS LVLAK---LG
SARS SSSRSRGN- - - - - - - SRNST PGSSRGNS- - - - - - - PARMA SGGGETA.A. LLDRLNOLE
ABW -RSTAASS- - - - - -AAASRA PSREGSRG- - - - - - - RRSDS GDDIARAAK IICD-----
... .... . .. .. ..... . . ... .... . . ... ..... . .... .. . .. . . ... .. .. . .

305 315 325 335 345 355
EMCR SPSSSGTSTP K- - - - - - - K- - - -PNKPLSO, PRADKPS- - - -OLKKPRWKR WPTR--EENV
229E DKKSAKTGTP KPSRNOSPAS SOTSAKSLAR SQSSETKEQK HEMQKPRWKR QPNDDVTSNV
PEOW HKQQQKPKQE K-SDN---SG KNTPKKNKSR ATSKERD- - - -LKDIPEWRR IPKG--ENSW
TGEW QRSRSKSKER S--------- - - ---NSKTR TTPKNE- - - - - -NRHTWKR AGK---GDW
FeCoW -RSRSKPRER S- - r - - - - - - - - - - - DSKR dTTPKNA- - - - - - NKHTWKK TAGE---GDW
PRCoW QRSRSKSKER S-- - - - - - - - - - - - -NSKTR DTTPKNE- - - ---NKHTWKR TAGK---GDV
CaCOW -RSRSKSKERS- - - - - - - - - - - - - - SSKTR DTTPKNE- - - - - - NKHWKR TAGE---GDW
RSDACoW -KDAGQPKQV Ta- - - - - - - - - - - - - KOSAK EVROKIL- - - - - -NKPROKR TPNK--CCPV
MW -KDAGQPKQV T-- - - - - - - - - - - - - KOSAK EVROK - - - - - -TKPRQKR, TPNK--QCPW
PHEW -KDATKPQOW T--------- - - - - - KOTAK EVROKIL- - - - - -NKPRQKR SPNK--QCTV
OC43 -KDATKPOV T- - - - - - - - - - - - - - KHTAK EVROKIL- - - - - -NKPROKR SPNK--CCTV
BoCoW -KDATKPOOV T--------a - - - - - KOTAK EIROKIL- - - - - - NKPROKR SPNK--CCTV
SARS SKVSGKGOOO O-a- - - - - - - - as a -GOTWTK KSAAEAS- - - - - - KKPROKR TATK--QYNV
ABW ---OKKGSRI T- - - - - - - - - - - - - -KAKAD EMAHRRY- - - - - - - - -CKRT IPPN---YRW
.. .. ..... . . ... ... . . ... . .. . .. . ... . . .... . . . .. .... . .... . ....
355 375 385 395 405 415
EMCR IOCFGPRDFN H---NMGDSD LVONGWDAKG FPOLAELIPN OAAIFFDSEV STDEVG----
229E TQCFGPRDLD H---NFGSAG WVANGVKAKG YPQFAELVPS TAAMLFDSHI VSKESG----
PEDW AACFGFRGGF K---NFGDAE, EVEKGWDASG YAQIASLAPN WAALLFGGNV AVRELA-----
TGEW TRFYGARSSS A---NFGDTD LVANGSSAKH YPOLAECVPS VSSILFGSYW TSKEDG----
FeCoW TTFYGARSSS A---NFGDSD LVANGNAAKC YPOIAECVPS VSSIIFGSQW SAEEAG----
PRCOW TRFYGARSSS A---NFGDSD LVANGSSAKH YPOLAECVPS WSSILFGSYW TSKEDG-- as
CaCoW TKFYGARSSS A---NFGDSD LVANGNGAKH YPOLAECWES WSSILEGSHW TAKEDG----
RSDACOW QQCFGKRGPN Q---NFGGPE MLKLGTSDPO FPILAELAPT PGAFFFGSKL, ELVKKN--SG
MHW QQCFGKRGPN Q---NFGGSE MLKLGTSDPQ FPILAELAPT PSAFFFGSKL, ELVKKN--SG
PHEV QOCEGKRGPN ---NFGGGE MKLGSDPO FPILAELAPT AGAEFFGSRL, ELAKVONLSG
OC43 OOCFGKRGPN 0---NFGGGE MLKLGTSDPO FPILAELAPT AGAFFFGSRL ELAKVONLSG
BoCoW OOCFGKRGPN Q---NFGGGE MLKLGTSDPO FPILAELAPT AGAFFFGSRL ELAKVONLSG
SARS TQAFGRRGPE OTQGNFGDQD LIRCGTDYKH WPQIAQFAPS ASAFFGMSRI GMEVTP----
ABW DOWFGPRTKG K-EGNFGDDK MNEEGIKDGR WTAMLNLVPS SHACLEGSRV TPKLOL----
... ..... . .... ..... . . ... ..... . .... ..... .... ..... . .... .. ..
A25 435 445 4S5 465 475
EMCR - - - - - - - DNV OITYT----Y KMLVAKDNKN LPKFIEOISA FTKPS----- SIKEMOSQSS
229E - - as a s is a NTV WLTFT----T RWTWPKDHH LGKFLEELNA FTR- - - - - - - EMOOHPLLNP
PEDW - - - - - - - DSY ETYN----Y KMTWPKSDPN WELLWSOVDA FKTGN- - - - - AKLQRKKEKK
TGEW - - - - - - - DOI EVTET----H KYHLPKDDPK TGOFLOOINA YARPS ----- EVAKEORKRK
FeCoW -- as as - - - Dov KVTLT----H TYYLPKDDAK TSOFLEQIDA YKRPS----- EVAKDORQRR
PRCOW - - - - - - - DQI EVTET----H KYHLPKDHPK TEQFLQQINA YASPs----- EAKEORKRK
CaCOW - - - - - - - DOI EWTE----H KYHLPKDdPK TGOFOONA YARPS- - - - - EVAKEORORK
RSDACW GVDEPTKDVY ELOYSGAVRF DSTLPGFETE MKVLNENLNA YONOA----- GGADVVSPKP
MHW GADEPTKDVY ELOYSGAIRF DSTLPGFETI MKVTENLNA Y-DOA- - - - - GSWDWSPKP
PHEV NPDEPQKDVY ELRYNGAIRF DSTLSGFETI MKVLNQNLNA YOHOE----- DGMMNISPKP
OC43 NPDEPOKDVY ELRYNGAIRE DSTLSGFETI MKVLNENLNA YQQQ------ DGMMNMSPKP
BoCoW NLDEPOKDVY ELRYNGAIRF DSTLSGFETI MKVLNENLNA YOOO------ DGMMNMSPKP
SARS - a-- - - - - SGT WLTYHGAIKL DDKDPOFKDN VILLNKHIDA YKTFP----- PTEPKKKKK
ABV - - - - - - - DGL HLRFEFTTVW PCDDPQFDNY WKICDQCVDG VGTRPKDDEP KPKSRSSSRP
.. . ..... ... ..... . ... ..... .

485 495 505 515 525
EMCR HVAQNTVLN- ---------- -ASPES - - - -KPLADDDSA EIWNEWLH
229E SALEFNPSQ- ---------- -TSPATA- - - -EPVRDEVSI EDIDEVN
PEOW NKRETTLQQH EEAIYDDVGA PSDWTHANLE WDTAWDGGDT AWEINEFD TGN- - - - - - -
TGEW SRSKSAERS- - - - - - - - - - - EODVWPDALI ENYTDVFDDT OVEIIDEVTN
FeCoW SRSKSADK- - - - - - - - - - - - KPEELSWTLV EAYTDWFDDT QVEMIDEVTN
PRCOW SRSKSAERS- - - - - - - - - - - EQEWVPDSLI ENYTDVFDDT OVEMIDEVTN
CaCoW ARSKSWERV- - - - - - - - - - - EQEWVPDALT ENYTDVFDDT QVEIIDEVTN
RSDACOW ORKRGTKQT- -AQKEELDSI SVAKPKSAVQ RNWSRELTPE DRSLLAQILD DGWVPDGLDD
MHV PRRGRRQAQ- -EKKDEVDNV SVAKPKSLVO RNWSRELTPE DRSLLAOILD DGVWPDGLED
PHEV ORQRGOKN- - --GOVENDNV SWAAPKSRVO CNKSRELTAE DISLLKKMDE P- - - - - YTED
OC43 QRQRGHKN- - --GQGENDNI SWAVPKSRVQ QNKSRELTAE DISLLKKMDE Pa- - - - - YTED
BoCoW QRORGQKN-- r-GOGENDNI SWAAPKSRVO ONKSRELTAE DSLLKKMOE P- - - - - YTED
SARS KTDEAQPLP- - - - - - - - - - - QRQKKOPTVT PAADMDDF SRQLONSMSG ASADSTO.A.--
AIBW ATRGNSPAPR QQRPKKEKKL. KKQDDEADKA LTSDEERNNA QLEFYDEPKy NWGOAAGE
EMCR
229E
PEOW
TGEW
FeCOW
PRCOW
CaCOV
RSDACOW -SNW
MHV DSNW
PHEW TSE
OC43 TSEI
BoCOW TSE
SARS
ABW NEL
k. 5'untranslated region (genomic sequence)

.... ...... .... ..... . .... ..... . .... ..... . .. ..... . .. .... .
5 5 25 55 35 45
EMCR5' UTR - p m - - - GAATTTTCTT ATTTAGACTT TGTGTCTACT
- -- - - - - -n a mass ps AGATAGA
229E5' UTR ACTTAAGTAC CTTATCTATC TACAGATAGA AAAGTTGCTT -TTTAGACTT TGTGTCTACT
.... ...... .... ......... .... . .... . .... . . .. . .... . .. . . .... .

65 75 85 95 105 115
EMCR5' UTR CCTCTCAACT AAACGAAATT TTT-CTAGTG CTGTCATTTG TTATG--GCA GTCCTAGTGT
229E5' UTR TTTCTCAACT AAACGAAATT TTTGCTATGG CCGGCATCTT TGATGCTGGA GTCGTAGTGT
... .....)
.. ..... .. ...... .. .. ..... . ... .... .
125 1.35 4S 155 1.65 l75
EMCR5' UTR AATTGAAATT TCGTCAAGTT TGTAA-ACTG GTTAGGCAAG TGTTGTATTT TCTGTGTTTA
229E5' UTR AATGAAATT TCATTTGGGT TGCAACAGTT TGGAAGCAAG TGCTGTGTGT CCTA-GTCTA
.. ..... .
... ...... .. .... . .... ..... .. ..... . ... . .... .
185 195 205 215 225 235
EMCR5' UTR AGCACTGGTG GTTCTGTC-C ACTAGTGCAC AC-ATTGATA CTTAAGT-GG TGTTCTGTCA
229E5' UTR AGGGTTTCGT GTTCCGTCAC GAGATTCCAT TCTACAAACG CCTTACTCGA GGTTCCGTCT
... ...... .. .... . .. .....) .. ..... . .. ..... .

245 255 26S 275 285
EMCRS' UTR CTGCTTATTG TGGAAGCAAC GTTCTGTCGT TGTGGAAACC AATAACGCT AACC
229E5' UTR CGTGTTTGTG TGGAAGCAAA GTTCTGTCTT TGTGGAAACC AGTAACTGTT CCTA
US 2008/0044426 A1 Feb. 21, 2008
NOVELATYPCAL PNEUMONA-CAUSING each isolate may have a somewhat different percentage

VIRUS relationship with the sequences of the isolate as provided
herein.
0008. When one wishes to compare a virus isolate with
The invention relates to the field of Virology. the sequences as listed in FIG. 1 molo, the invention
0001 provides an isolated essentially mammalian positive-sense
0002 Recently, a respiratory illness (atypical pneumo single stranded RNA virus (EMCR-CoV) belonging to the
nia) was diagnosed in an 8 months old patient that could not Coronaviruses and identifiable as phylogenetically corre
be attributed to SARS (Severe Acute Respiratory Syndrome) sponding thereto by determining a nucleic acid sequence of
virus or any other known viral infection. The patient tested said virus and determining that said nucleic acid sequence
negative for influenza, parainfluenza, mumps and RSV and has a percentage nucleic acid identity to the sequences as
yet the disease was identified to be caused by a virus which listed higher than the percentages identified herein for the
closely resembled SARS. nucleic acids as identified herein below in comparison with
0003 For being able to trace its origin, monitor its PEDV, 229E, PRCoV, TGEV, CaCoV and FeCoV. Likewise,
epidemiology and prevent possible spreading of the disease, an isolated essentially mammalian positive-sense single
it is of great importance to be able to recognise viral causes stranded RNA virus (EMCR-CoV) belonging to the Coro
of pneumonia in an early stage. Especially, if severe diseases naviruses and identifiable as phylogenetically corresponding
thereto by determining an amino acid sequence of said virus
are found to be caused by viruses, it is necessary to detect and determining that said amino acid sequence has a per
the identity of the virus as soon as possible, in order to centage amino acid homology to the sequences as listed
develop diagnostic tools and possibly therapies. The SARS which is essentially higher than the percentages provided
epidemy has shown that it is paramount for prevention of herein in comparison with PEDV, 229E, PRCoV, TGEV.
spread of the disease to be able to get an early diagnosis in CaCoV and FeCoV.
order to timely take effective isolation measures en initiate 0009. With the provision of the sequence information of
quarantaine precautions. Only then, world-wide contamina this EMCR-Coronavirus (EMCR-CoV), the invention pro
tions can be prevented. vides diagnostic means and methods, prophylactic means
0004 Furthermore, identification of the viral cause for and methods and therapeutic means and methods to be
the disease enables development of vaccines, which can be employed in the diagnosis, prevention and/or treatment of
used prophylactically to protect people who are at risk of disease, in particular of respiratory disease (atypical pneu
being infected. And, finally, knowledge of the viral cause monia), in particular of mammals, more in particular in
enables to develop therapeutic measures. humans associated with infection by this virus. In Virology,
0005 Thus, there is great need in developing diagnostic it is most advisory that diagnosis, prophylaxis and/or treat
tools and therapies for viral pneumonias in general, and ment of a specific viral infection is performed with reagents
particular to a novel disease-causing infectious agent, espe that are most specific for said specific virus causing said
cially when this agent appears to be a virus. infection. In this case this means that it is preferred that said
0006. The invention provides the nucleotide sequence of diagnosis, prophylaxis and/or treatment of an EMCR-CoV
an isolated essentially mammalian positive-sense single virus infection is performed with reagents that are most
stranded RNA virus belonging to the Coronaviruses, which specific for EMCR-CoV virus. This by no means however
is the causative factor for the new disease, hereinafter excludes the possibility that less specific, but sufficiently
referred to as EMCR-CoV and the disease being referred to cross-reactive reagents are used instead, for example
as EMCR-CoV-caused pneumonia. From a phylogenetic because they are more easily available and sufficiently
analysis of the Matrix and Nucleocapsid gene sequences of address the task at hand.
the virus (FIGS. 2a and 2b) it appears that the virus is a 0010. The invention for example provides a method for
distinct member of the group formed by PEDV (porcine Virologically diagnosing an EMCR-CoV infection of an
epidemic diarrhea virus), HCoV-229E (human coronavirus animal, in particular of a mammal, more in particular of a
229E), PRCoV (porcine respiratory coronavirus), TGEV human being, comprising determining in a sample of said
(transmissible gastroenteritis virus), CaCoV (Canine coro animal the presence of a viral isolate or component thereof
navirus) and FeCoV (feline coronavirus). Based on amino by reacting said sample with an EMCR-CoV specific nucleic
acid identity matrices, human coronavirus 229E seems to be acid or antibody according to the invention, and a method
the closest relative (for all ORFs with the exception of for serologically diagnosing an EMCR-CoV infection of a
Matrix which appears to be slightly more closely related to mammal comprising determining in a sample of said mam
PEDV see FIG. 3). mal the presence of an antibody specifically directed against
0007 Although phylogenetic analyses provide a conve an EMCR-CoV virus or component thereof by reacting said
nient method of identifying a virus, several other possibly sample with an EMCR-CoV virus-specific proteinaceous
more straightforward albeit somewhat more coarse methods molecule or fragment thereof or an antigen according to the
for identifying said virus or viral proteins or nucleic acids invention.
from said virus are herein also provided. As a rule of thumb 0011. The invention also provides a diagnostic kit for
an EMCR-Coronavirus can be identified by the percentages diagnosing an EMCR-CoV infection comprising an EMCR
of homology of the virus, proteins or nucleic acids to be CoV virus, an EMCR-CoV virus-specific nucleic acid, pro
identified in comparison with viral proteins or nucleic acids teinaceous molecule or fragment thereof, antigen and/or an
identified herein by sequence. It is generally known that antibody according to the invention, and preferably a means
virus species, especially RNA virus species, often constitute for detecting said EMCR-CoV virus, EMCR-CoV virus
a quasi species wherein a cluster of said viruses displays specific nucleic acid, proteinaceous molecule or fragment
heterogeneity among its members. Thus it is expected that thereof, antigen and/or an antibody, said means for example
US 2008/0044426 A1 Feb. 21, 2008
comprising an excitable group Such as a fluorophore or amino acid sequence identity, between ORFs of EMCR
enzymatic detection system used in the art (examples of CoV virus and any of the known other viruses of the same
suitable diagnostic kit format comprise IF ELISA, neutral family to date are with human coronavirus 299E or Porcine
ization assay, RT-PCR assay). To determine whether an as Epidemic Diarrhea Virus (see FIGS. 3 and 4). The amino
yet unidentified virus component or synthetic analogue acid identities with human coronavirus 229E ranges from
thereof Such as nucleic acid, proteinaceous molecule or 45% (Nucleoprotein) to 81% (Replicase 1b); interestingly,
fragment thereof can be identified as EMCR-CoV-virus Replicase 1a has an identity of just 56% contrasting with
specific, it suffices to analyse the nucleic acid or amino acid Replicase 1b's 81% identity. EMCR CoV has a closer
sequence of said component, for example for a stretch of identity with human coronavirus 229E than with any of the
said nucleic acid or amino acid, preferably of at least 10, known other viruses of the same family to date for all
more preferably at least 25, more preferably at least 40 putative ORFs, with the exception of Matrix, which is
nucleotides or amino acids (respectively), by sequence slightly more closely related to the Matrix ORF of PEDV.
homology comparison with the provided EMCR-CoV viral Individual proteins or whole virus isolates with, respec
sequences and with known non-EMCR-CoV viral sequences tively, higher homology than these mentioned maximum
(human coronavirus 299E is preferably used) using for values are considered phylogenetically corresponding and
example phylogenetic analyses as provided herein. Depend thus taxonomically corresponding to EMCR-CoV virus, and
ing on the degree of relationship with said EMCR-CoV or generally will be encoded by a nucleic acid sequence
non-EMCR-CoV viral sequences, the component or syn structurally corresponding with a sequence as shown in FIG.
thetic analogue can be identified. 1. Herewith the invention provides a virus phylogenetically
0012. The invention thus provides the nucleotide corresponding to the isolated virus of which the sequences
sequence of a novel etiological agent, an isolated essentially are depicted in FIG. 1.
mammalian positive-sense single stranded RNA virus 0015. It should be noted that, similar to other viruses, a
(herein also called EMCR-CoV virus) belonging to the certain degree of variation can be expected to be found
Coronaviridae family, and EMCR-CoV virus-specific com between EMCR-CoV-viruses isolated from different
ponents or synthetic analogues thereof. SOUCS.
0013 Coronaviruses were first isolated from chickens in (0016. Also, the viral sequence of the EMCR-CoV virus
1937, while the first human coronavirus was propagated in or an isolated EMCR-CoV virus gene as provided hereinfor
vitro by Tyrell and Bonoe in 1965. There are now about 13 example shows less than 95%, preferably less than 90%,
species in this family, which infect cattle, pigs, rodents, cats, more preferably less than 80%, more preferably less than
dogs, birds and man. Coronavirus particles are irregularly 70% and most preferably less than 65% nucleotide sequence
shaped, about 60-220 nm in diameter, with an outer enve homology or less than 95%, preferably less than 90%, more
lope bearing distinctive, club-shaped peplomers (about 20 preferably less than 80%, more preferably less than 70% and
nm long and 10 nm wide at the distal end). This crown-like most preferably less than 65% amino acid sequence homol
appearance give the family its name. The envelope carries ogy with the respective nucleotide or amino acid sequence
two glycoproteins: S, the spike glycoprotein which is of the human coronavirus 299E or Porcine Epidemic Diar
involved in cell fusion and is a major antigen, and M, the rhea Virus as for example can be found in Genbank (for
membrane glycoprotein, which is involved in budding and example in accession number af304460 (HCoV-299E) or
envelope formation. The genome is associated with a basic af353511 (PEDV).
phosphoprotein, designated N. The genome of coronavi (0017 Sequence divergence of EMCR-CoV strains
ruses, a single stranded positive-sense RNA strand, is typi around the world may be somewhat higher, in analogy with
cally 27-31 Kb long and contains a 5' methylated cap and a other coronaviruses.
3' poly-A tail, by which it can directly function as an mRNA 0018. The term “nucleotide sequence homology” as used
in the infected cell. Initially the 5' ORF 1 (about 20 Kb) is herein denotes the presence of homology between two
translated to produce a viral polymerase, which then pro (poly)nucleotides. Polynucleotides have “homologous'
duces a full length negative sense Strand. This is used as a sequences if the sequence of nucleotides in the two
template to produce mRNA as a nested set of transcripts, sequences is the same when aligned for maximum corre
all with identical 5' non-translated leader sequence of 72 spondence. Sequence comparison between two or more
nucleotides and coincident 3' polyadenylated ends. Each polynucleotides is generally performed by comparing por
mRNA thus produced is monocistronic, the genes at the 5' tions of the two sequences over a comparison window to
end being translated from the longest mRNA and so on. identify and compare local regions of sequence similarity.
These unusual cytoplasmic structures are produced not by The comparison window is generally from about 20 to 200
splicing, but by the polymerase during transcription. contiguous nucleotides. The “percentage of sequence
Between each of the genes there is a repeated intergenic homology' for polynucleotides, such as 50, 60, 70, 80, 90,
sequence AACUAAAC which interacts with the tran 95, 98, 99 or 100 percent sequence homology may be
Scriptase plus cellular factors to splice the leader sequence determined by comparing two optimally aligned sequences
onto the start of each ORF. In some coronaviruses there are over a comparison window, wherein the portion of the
about 8 ORFs, coding for the proteins mentioned above, but polynucleotide sequence in the comparison window may
also for a heamagglutenin esterase (HE), and several other include additions or deletions (i.e. gaps) as compared to the
non-structural proteins. reference sequence (which does not comprise additions or
0014 Newly isolated viruses are phylogenetically corre deletions) for optimal alignment of the two sequences. The
sponding to and thus taxonomically corresponding to percentage is calculated by: (a) determining the number of
EMCR-CoV virus when comprising a gene order and/or positions at which the identical nucleic acid base occurs in
amino acid sequence and/or nucleotide sequence Sufficiently both sequences to yield the number of matched positions; (b)
similar to our prototypic EMCR-CoV virus. The highest dividing the number of matched positions by the total
US 2008/0044426 A1 Feb. 21, 2008
number of positions in the window of comparison; and (c) depend on many factors, including temperature and source
multiplying the result by 100 to yield the percentage of of primer. A “pair of bidirectional primers' as used herein
sequence homology. Optimal alignment of sequences for refers to one forward and one reverse primer as commonly
comparison may be conducted by computerized implemen used in the art of DNA amplification such as in PCR
tations of known algorithms, or by inspection. Readily amplification.
available sequence comparison and multiple sequence align 0023 The term “probe' refers to a single-stranded oli
ment algorithms are, respectively, the Basic Local Align gonucleotide sequence that will recognize and form a hydro
ment Search Tool (BLAST) (Altschul, S. F. et al. 1990. J. gen-bonded duplex with a complementary sequence in a
Mol. Biol. 215:403; Altschul, S. F. et al. 1997. Nucleic Acid target nucleic acid sequence analyte or its cDNA derivative.
Res. 25:3389-3402) and ClustalW programs both available 0024. The terms “stringency” or “stringent hybridization
on the internet. Other suitable programs include GAP, conditions’ refer to hybridization conditions that affect the
BESTFIT and FASTA in the Wisconsin Genetics Software stability of hybrids, e.g., temperature, Salt concentration, pH,
Package (Genetics Computer Group (GCG), Madison, Wis., formamide concentration and the like. These conditions are
USA). empirically optimised to maximize specific binding and
0019. As used herein, “substantially complementary’ minimize non-specific binding of primer or probe to its
means that two nucleic acid sequences have at least about target nucleic acid sequence. The terms as used include
65%, preferably about 70%, more preferably about 80%, reference to conditions under which a probe or primer will
even more preferably 90%, and most preferably about 98%, hybridise to its target sequence, to a detectably greater
sequence complementarity to each other. This means that the degree than other sequences (e.g. at least 2-fold over back
primers and probes must exhibit Sufficient complementarity ground). Stringent conditions are sequence dependent and
to their template and target nucleic acid, respectively, to will be different in different circumstances. Longer
hybridise under stringent conditions. Therefore, the primer sequences hybridise specifically at higher temperatures.
sequences as disclosed in this specification need not reflect Generally, stringent conditions are selected to be about 5°C.
the exact sequence of the binding region on the template and lower than the thermal melting point (Tm) for the specific
degenerate primers can be used. A Substantially complemen sequence at a defined ionic strength and pH. The Tm is the
tary primer sequence is one that has sufficient sequence temperature (under defined ionic strength and pH) at which
complementarity to the amplification template to result in 50% of a complementary target sequence hybridises to a
primer binding and second-strand synthesis. perfectly matched probe or primer. Typically, stringent con
0020. The term “hybrid” refers to a double-stranded ditions will be those in which the salt concentration is less
nucleic acid molecule, or duplex, formed by hydrogen than about 1.0 M Na+ion, typically about 0.01 to 1.0 M
bonding between complementary nucleotides. The terms Na+ion concentration (or other salts) at pH 7.0 to 8.3 and the
“hybridise” or “anneal refer to the process by which single temperature is at least about 30° C. for short probes or
Strands of nucleic acid sequences form double-helical seg primers (e.g. 10 to 50 nucleotides) and at least about 60° C.
ments through hydrogen bonding between complementary for long probes or primers (e.g. greater than 50 nucleotides).
nucleotides. Stringent conditions may also be achieved with the addition
0021. The term "oligonucleotide' refers to a short of destabilizing agents such as formamide. Exemplary low
sequence of nucleotide monomers (usually 6 to 100 nucle stringent conditions or “conditions of reduced stringency’
otides) joined by phosphorous linkages (e.g., phosphodi include hybridization with a buffer solution of 30% forma
ester, alkyl and aryl-phosphate, phosphorothioate), or non mide, 1 M NaCl, 1% SDS at 37° C. and a wash in 2XSSC
phosphorous linkages (e.g., peptide, Sulfamate and others). at 40° C. Exemplary high Stringency conditions include
An oligonucleotide may contain modified nucleotides hav hybridization in 50% formamide, 1 MNaCl, 1% SDS at 37°
ing modified bases (e.g., 5-methyl cytosine) and modified C., and a wash in 0.1xSSC at 60° C. Hybridization proce
Sugar groups (e.g. 2'-O-methyl ribosyl, 2'-O-methoxyethyl dures are well known in the art and are described in e.g.
ribosyl, 2'-fluoro ribosyl, 2-amino ribosyl, and the like). Ausubel et al. Current Protocols in Molecular Biology, John
Oligonucleotides may be naturally-occurring or synthetic Wiley & Sons Inc., 1994.
molecules of double- and single-stranded DNA and double 0025. The term “antibody' includes reference to antigen
and single-stranded RNA with circular, branched or linear binding forms of antibodies (e.g., Fab, F (ab) 2). The term
shapes and optionally including domains capable of forming “antibody' frequently refers to a polypeptide substantially
stable secondary structures (e.g., stem-and-loop and loop encoded by an immunoglobulin gene or immunoglobulin
stem-loop structures). genes, or fragments thereof which specifically bind and
0022. The term “primer' as used herein refers to an recognize an analyte (antigen). However, while various
oligonucleotide which is capable of annealing to the ampli antibody fragments can be defined in terms of the digestion
fication target allowing a DNA polymerase to attach thereby of an intact antibody, one of skill will appreciate that such
serving as a point of initiation of DNA synthesis when fragments may be synthesized de novo either chemically or
placed under conditions in which synthesis of primer exten by utilizing recombinant DNA methodology. Thus, the term
sion product which is complementary to a nucleic acid antibody, as used herein, also includes antibody fragments
Strand is induced, i.e., in the presence of nucleotides and an Such as single chain Fv, chimeric antibodies (i.e., compris
agent for polymerization Such as DNA polymerase and at a ing constant and variable regions from different species),
suitable temperature and pH. The (amplification) primer is humanized antibodies (i.e., comprising a complementarity
preferably single stranded for maximum efficiency in ampli determining region (CDR) from a non-human source) and
fication. Preferably, the primer is an oligodeoxyribonucle heteroconjugate antibodies (e.g., bispecific antibodies).
otide. The primer must be sufficiently long to prime the 0026. In short, the invention provides an isolated essen
synthesis of extension products in the presence of the agent tially mammalian positive-sense single stranded RNA virus
for polymerization. The exact lengths of the primers will (EMCR-CoV) belonging to the Coronaviruses and identifi
US 2008/0044426 A1 Feb. 21, 2008
able as phylogenetically corresponding thereto by determin of the analysed S-protein encoded by a sequence comprising
ing a nucleic acid sequence of a Suitable fragment of the the sequence of translation 2 of E and translation 1 of the F
genome of said virus and testing it in phylogenetic tree sequence of the S-protein as depicted in FIG. 1 is found, the
analyses wherein maximum likelihood trees are generated analysed virus isolate comprises an EMCR-CoV virus iso
using 100 bootstraps and 3 jumbles and finding it to be more late according to the invention. The S ORF of the EMCR
closely phylogenetically corresponding to a virus isolate CoV virus seems to be located adjacent to the ORF lab
having the sequences as depicted in FIG. 1 than it is (coding for the viral replicase), which would discriminate an
corresponding to a virus isolate of PEDV (porcine epidemic EMCR-CoV viruses from the bovine coronavirus and the
diarrhea virus), HCoV-229E (human coronavirus 229E), murine hepatitis virus, which have a so-called 2a gene and
PRCoV (porcine respiratory coronavirus), TGEV (transmis an HE-gene between the S protein and the viral polymerase.
sible gastroenteritis virus), CaCoV (Canine coronavirus) and 0033. The invention provides among others an isolated or
FeCoV (feline coronavirus). recombinant nucleic acid or virus-specific functional frag
0027 Suitable nucleic acid genome fragments each use ment thereof obtainable from a virus according to the
ful for Such phylogenetic tree analyses are for example any invention. The isolated or recombinant nucleic acids com
of the fragments encoding the Matrix protein or the Nucleo prises the sequences as given in FIG. 1 or sequences of
capsid protein as disclosed in FIG. 1, leading to the phylo homologues which are able to hybridise with those under
genetic tree analysis as disclosed herein in FIG. 2a or 2b. stringent conditions. In particular, the invention provides
Other suitable nucleic acid fragments useful for such phy primers and/or probes suitable for identifying an EMCR
logenetic tree analyses are for example any of the fragments CoV virus nucleic acid.
encoding Replicase 1a and 1b, Spike, orf 4a and 4b, and E. 0034) Furthermore, the invention provides a vector com
0028. A suitable open reading frame (ORF) useful in prising a nucleic acid according to the invention. To begin
phylogenetic analyses comprises the ORF encoding the viral with, vectors such as plasmid vectors containing (parts of)
replicase (ORF1a). When an overall amino acid identity of the genome of the EMCR-CoV virus, virus vectors contain
at least 60%, preferably of at least 70%, more preferably of ing (parts of) the genome of the EMCR-CoV (for example,
at least 80%, more preferably of at least 90%, most prefer but not limited thereto, vaccinia virus, retroviruses, bacu
ably of at least 95% of the analysed replicase with the lovirus), or EMCR-CoV virus containing (parts of) the
replicase having a sequence comprising the amino acids of genome of other viruse or other pathogens are provided.
FIG. 1 is found, the analysed virus isolate comprises an 0035 Also, the invention provides a host cell comprising
EMCR-CoV virus isolate according to the invention. a nucleic acid or a vector according to the invention. Plasmid
0029. A suitable open reading frame (ORF) useful in or viral vectors containing the replicase components of
phylogenetic analyses comprises the ORF encoding the viral EMCR-CoV virus are generated in prokaryotic cells for the
replicase (ORF 1b). When an overall amino acid identity of expression of the components in relevant cell types (bacte
at least 82%, more preferably of at least 90%, most prefer ria, insect cells, eukaryotic cells). Plasmid or viral vectors
ably of at least 95% of the analysed replicase with the containing full-length or partial copies of the EMCR-CoV
replicase having a sequence comprising the amino acids of virus genome will be generated in prokaryotic cells for the
FIG. 1 is found, the analysed virus isolate comprises an expression of viral nucleic acids in-vitro or in-vivo. The
EMCR-CoV virus isolate according to the invention. latter vectors may contain other viral sequences for the
0030. Another suitable open reading frame (ORF) useful generation of chimeric viruses or chimeric virus proteins,
in phylogenetic analyses comprises the ORF encoding the may lack parts of the viral genome for the generation of
Nucleocapsid protein. When an overall amino acid identity replication defective virus, and may contain mutations,
of at least 50%, more preferably of at least 60%, more deletions or insertions for the generation of attenuated
preferably of at least 70%, more preferably of at least 80%, viruses.
more preferably of at least 90%, most preferably of at least 0036) Infectious copies of EMCR-CoV virus (being wild
95% of the analysed Nucleocapsid protein with the Nucleo type, attenuated, replication-defective or chimeric) can be
capsid protein encoded by a sequence comprising (part of) produced upon co-expression of the polymerase components
the sequence F of FIG. 1 is found, the analysed virus isolate according to the state-of-the-art technologies described
comprises an EMCR-CoV isolate according to the inven above.
tion. 0037. In addition, eukaryotic cells, transiently or stably
0031. Another suitable open reading frame (ORF) useful expressing one or more full-length or partial EMCR-CoV
in phylogenetic analyses comprises the ORF encoding the virus proteins can be used. Such cells can be made by
Matrix protein. When an overall amino acid identity of at transfection (proteins or nucleic acid vectors), infection
least 60%, more preferably of at least 70%, more preferably (viral vectors) or transduction (viral vectors) and may be
of at least 80%, more preferably of at least 90%, most useful for complementation of mentioned wild type, attenu
preferably of at least 95% of the analysed Matrix protein ated, replication-defective or chimeric viruses.
with the Matrix protein encoded by a sequence comprising 0038 A chimeric virus may be of particular use for the
(part of) the sequence F of FIG. 1 is found, the analysed generation of recombinant vaccines protecting against two
virus isolate comprises an EMCR-CoV isolate according to or more viruses. For example, it can be envisaged that
the invention. EMCR-CoV virus vector expressing one or more proteins of
0032. Another suitable open reading frame (ORF) useful a human metapneumovirus or a human metapneumovirus
in phylogenetic analyses comprises the ORF encoding the vector expressing one or more proteins of EMCR-CoV virus
spike protein S. When an overall amino acid identity of at will protect individuals vaccinated with Such vector against
least 55%, more preferably of at least 60%, more preferably both virus infections. Such a specific chimeric virus is
of at least 70%, more preferably of at least 80%, more particularly useful in the invention because it is Suspected
preferably of at least 90%, most preferably of at least 95% that co-infection of for instance, human metapneumovirus
US 2008/0044426 A1 Feb. 21, 2008
frequently occurs in coronavirus infected patients. Attenu such as the Polymerase Chain Reaction (PCR; Mullis 1987,
ated and replication-defective viruses may be of use for U.S. Pat. Nos. 4,683,195, 4,683.202, en 4,800,159) or by
vaccination purposes with live vaccines as has been Sug using amplification reactions such as Ligase Chain Reaction
gested for other viruses. (LCR; Barany 1991, Proc. Natl. Acad. Sci. USA 88:189
0039. In a preferred embodiment, the invention provides 193: EP Appl. No. 320,308), Self-Sustained Sequence Rep
a proteinaceous molecule or coronavirus-specific viral pro lication (3SR: Guatelli et al., 1990, Proc. Natl. Acad. Sci.
tein or functional fragment thereof encoded by a nucleic acid USA 87: 1874-1878), Strand Displacement Amplification
according to the invention. Useful proteinaceous molecules (SDA: U.S. Pat. Nos. 5,270,184, en 5,455,166), Transcrip
are for example derived from any of the genes or genomic tional Amplification System (TAS; Kwoh et al., Proc. Natl.
fragments derivable from a virus according to the invention. Acad. Sci. USA 86:1173-1177), Q-Beta Replicase (IAZardi
Such molecules, or antigenic fragments thereof, as provided et al., 1988, Bio/Technology 6:1197), Rolling Circle Ampli
herein, are for example useful in diagnostic methods or kits fication (RCA; U.S. Pat. No. 5,871,921), Nucleic Acid
and in pharmaceutical compositions such as Sub-unit vac Sequence Based Amplification (NASBA), Cleavase Frag
cines and inhibitory peptides. Particularly useful are the ment Length Polymorphism (U.S. Pat. No. 5,719,028),
viral replicase protein, the spike protein, the matrix protein,
the nucleocapsid orantigenic fragments thereof for inclusion Isothermal and Chimeric Primer-initiated Amplification of
as antigen or subunit immunogen, but inactivated whole Nucleic Acid (ICAN), Ramification-extension Amplification
virus can also be used. Particularly useful are also those Method (RAM: U.S. Pat. Nos. 5,719,028 and 5,942,391) or
proteinaceous Substances that are encoded by recombinant other suitable methods for amplification of nucleic acids.
nucleic acid fragments that are identified for phylogenetic 0045. In order to amplify a nucleic acid with a small
analyses, of course preferred are those that are within the number of mismatches to one or more of the amplification
preferred bounds and metes of ORFs useful in phylogenetic primers, an amplification reaction may be performed under
analyses, in particular for eliciting EMCR-CoV virus spe conditions of reduced stringency (e.g. a PCR amplification
cific antibodies, whether in vivo (e.g. for protective puposes using an annealing temperature of 38°C., or the presence of
or for providing diagnostic antibodies) or in vitro (e.g. by 3.5 mM MgCl2). The person skilled in the art will be able
phage display technology or another technique useful for to select conditions of Suitable stringency.
generating synthetic antibodies). 0046. The primers herein are selected to be “substan
0040 Also provided herein are antibodies, be it natural tially” complementary (i.e. at least 65%, more preferably at
polyclonal or monoclonal, or synthetic (e.g. (phage) library least 80% perfectly complementary) to their target regions
derived binding molecules) antibodies that specifically react present on the different Strands of each specific sequence to
with an antigen comprising a proteinaceous molecule or be amplified. It is possible to use primer sequences contain
EMCR-CoV virus-specific functional fragment thereof ing e.g. inositol residues or ambiguous bases or even primers
according to the invention. Such antibodies are useful in a that contain one or more mismatches when compared to the
method for identifying a viral isolate as an EMCR-CoV target sequence. In general, sequences that exhibit at least
virus comprising reacting said viral isolate or a component 65%, more preferably at least 80% homology with the target
thereof with an antibody as provided herein. This can for DNA or RNA oligonucleotide sequences, are considered
example be achieved by using purified or non-purified suitable for use in a method of the present invention.
EMCR-CoV virus or parts thereof (proteins, peptides) using Sequence mismatches are also not critical when using low
ELISA, RIA, FACS or similar formats of antigen detection stringency hybridization conditions.
assays (Current Protocols in Immunology). Alternatively, 0047. The detection of the amplification products can in
infected cells or cell cultures may be used to identify viral principle be accomplished by any suitable method known in
antigens using classical immunofluorescence or immunohis the art. The detection fragments may be directly stained or
tochemical techniques. Specifically useful in this respect are labelled with radioactive labels, antibodies, luminescent
antibodies raised against EMCR-CoV virus proteins which dyes, fluorescent dyes, or enzyme reagents. Direct DNA
are encoded by a nucleotide sequence comprising one or stains include for example intercalating dyes such as acri
more of the sequences disclosed in FIG. 1. dine orange, ethidium bromide, ethidium monoaZide or
0041. Other methods for identifying a viral isolate as an Hoechst dyes. Alternatively, the DNA or RNA fragments
EMCR-CoV virus comprise reacting said viral isolate or a may be detected by incorporation of labelled dNTP bases
component thereof with a virus specific nucleic acid accord into the synthesized fragments. Detection labels which may
ing to the invention. be associated with nucleotide bases include e.g. fluorescein,
0042. In this way the invention provides a viral isolate cyanine dye or BrdUrd.
identifiable with a method according to the invention as a 0048. When using a probe-based detection system, a
mammalian virus taxonomically corresponding to a posi suitable detection procedure for use in the present invention
tive-sense single stranded RNA virus identifiable as likely may for example comprise an enzyme immunoassay (EIA)
belonging to the EMCR-CoV virus genus within the family format (Jacobs et al., 1997, J. Clin. Microbiol. 35,791-795).
of Coronaviruses. For performing a detection by manner of the EIA procedure,
0043. The method is useful in a method for virologically either the forward or the reverse primer used in the ampli
diagnosing an EMCR-CoV virus infection of a mammal, fication reaction may comprise a capturing group, such as a
said method for example comprising determining in a biotin group for immobilization of target DNA PCR ampli
sample of said mammal the presence of a viral isolate or cons on e.g. a streptavidin coated microtiter plate wells for
component thereof by reacting said sample with a nucleic subsequent EIA detection of target DNA -amplicons (see
acid or an antibody according to the invention. below). The skilled person will understand that other groups
0044) Methods of the invention can in principle be per for immobilization of target DNA PCR amplicons in an EIA
formed by using any nucleic acid amplification method, format may be employed.
US 2008/0044426 A1 Feb. 21, 2008
0049. Probes useful for the detection of the target DNA as gency conditions can utilize a hybridization and/or wash at
disclosed herein preferably bind only to at least a part of the 11, 12, 13, 14, 15, or 20° C. lower than the thermal melting
DNA sequence region as amplified by the DNA amplifica point (Tm). Using the equation, hybridization and wash
tion procedure. Those of skill in the art can prepare suitable compositions, and desired Tm, those of ordinary skill will
probes for detection based on the nucleotide sequence of the understand that variations in the Stringency of hybridization
target DNA without undue experimentation as set out herein. and/or wash solutions are inherently described. If the desired
Also the complementary nucleotide sequences, whether degree of mismatching results in a Tm of less than 45° C.
DNA or RNA or chemically synthesized analogs, of the (aqueous solution) or 32° C. (formamide solution) it is
target DNA may suitably be used as type-specific detection preferred to increase the SSC concentration so that a higher
probes in a method of the invention, provided that such a temperature can be used. An extensive guide to the hybrid
complementary strand is amplified in the amplification reac ization of nucleic acids is found in Tijssen, Laboratory
tion employed. Techniques in Biochemistm and Molecular Biology—Hy
0050 Suitable detection procedures for use herein may bridization with Nucleic Acid Probes, Part I, Chapter 2
for example comprise immobilization of the amplicons and “Overview of principles of hybridization and the strategy of
probing the DNA sequences thereof by e.g. southern blot nucleic acid probe assays, Elsevier. New York (1993); and
ting. Other formats may comprise an EIA format as Current Protocols in Molecular Biology, Chapter 2, Ausubel,
described above. To facilitate the detection of binding, the et al., Eds. Greene Publishing and Wiley-Interscience, New
specific amplicon detection probes may comprise a label York (1995).
moiety Such as a fluorophore, a chromophore, an enzyme or 0053. In another aspect, the invention provides oligo
a radio-label. So as to facilitate monitoring of binding of the nucleotide probes for the generic detection of target RNA or
probes to the reaction product of the amplification reaction. DNA. The detection probes herein are selected to be “sub
Such labels are well-known to those skilled in the art and stantially” complementary to one of the strands of the
include, for example, fluorescein isothiocyanate (FITC). double stranded nucleic acids generated by an amplification
B-galactosidase, horseradish peroxidase, Streptavidin, reaction of the invention. Preferably the probes are substan
biotin, digoxigenin, 35S or 125I. Other examples will be tially complementary to the immobilizable, e.g. biotin
apparent to those skilled in the art. labelled, antisense Strands of the amplicons generated from
0051 Detection may also be performed by a so called the target RNA or DNA.
reverse line blot (RLB) assay, such as for instance described 0054. It is allowable for detection probes of the present
by Van den Brule et al. (2002, J. Clin. Microbiol. 40, invention to contain one or more mismatches to their target
779-787). For this purpose RLB probes are preferably sequence. In general, sequences that exhibit at least 65%,
synthesized with a 5' amino group for Subsequent immobi more preferably at least 80% homology with the target
lization on e.g. carboxyl-coated nylon membranes. The oligonucleotide sequences are considered suitable for use in
advantage of an RLB format is the ease of the system and its a method of the present invention.
speed, thus allowing for high throughput sample processing. 0055 Antibodies, both monoclonal and polyclonal, can
0052. The use of nucleic acid probes for the detection of also be used for detection purpose in the present invention,
RNA or DNA fragments is well known in the art. Mostly for example, in immunoassays in which they can be utilized
these procedure comprise the hybridization of the target in liquid phase or bound to a solid phase carrier. In addition,
nucleic acid with the probe followed by post-hybridization the monoclonal antibodies in these immunoassays can be
washings. Specificity is typically the function of post-hy detectably labeled in various ways. A variety of immunoas
bridization washes, the critical factors being the ionic say formats may be used to select antibodies specifically
strength and temperature of the final wash solution. For reactive with a particular protein (or other analyte). For
nucleic acid hybrids, the Tm can be approximated from the example, Solid-phase ELISA immunoassays are routinely
equation of Meinkoth and Wahl, Anal. Biochem., 138: used to select monoclonal antibodies specifically immunore
267-284 (1984): Tm=81.5° C.+16.6(log M)+0.41(% GC)- active with a protein. See Harlow and Lane, Antibodies. A
0.61 (% form)-500/L: where M is the molarity of monova Laboratory Manual, Cold Spring Harbor Publications, New
lent cations, 96 GC is the percentage of guanosine and York (1988), for a description of immunoassay formats and
cytosine nucleotides in the nucleic acid, '% form is the conditions that can be used to determine selective binding.
percentage of formamide in the hybridization Solution, and Examples of types of immunoassays that can utilize anti
L is the length of the hybrid in base pairs. The Tm is the bodies of the invention are competitive and non-competitive
temperature (under defined ionic strength and pH) at which immunoassays in either a direct or indirect format.
50% of a complementary target sequence hybridizes to a Examples of such immunoassays are the radioimmunoassay
perfectly matched probe. Tm is reduced by about 1° C. for (RIA) and the Sandwich (immunometric) assay. Detection of
each 1% of mismatching; thus, the hybridization and/or the antigens using the antibodies of the invention can be
wash conditions can be adjusted to hybridize to sequences of done utilizing immunoassays that are run in either the
the desired identity. For example, if sequences with >90% forward, reverse, or simultaneous modes, including immu
identity are sought, the Tm can be decreased 10° C. Gen nohistochemical assays on physiological samples. Those of
erally, stringent conditions are selected to be about 5° C. skill in the art will know, or can readily discern, other
lower than the thermal melting point (Tm) for the specific immunoassay formats without undue experimentation.
sequence and its complement at a defined ionic strength and 0056 Antibodies can be bound to many different carriers
pH. However, severely stringent conditions can utilize a and used to detect the presence of the target molecules.
hybridization and/or wash at 1, 2, 3, or 4°C. lower than the Examples of well-known carriers include glass, polystyrene,
thermal melting point (Tm); moderately stringent conditions polypropylene, polyethylene, dextran, nylon, amylases,
can utilize a hybridization and/or wash at 6, 7, 8, 9, or 10° natural and modified celluloses, polyacrylamides, agaroses
C. lower than the thermal melting point (Tm): low strin and magnetite. The nature of the carrier can be either soluble
US 2008/0044426 A1 Feb. 21, 2008
or insoluble for purposes of the invention. Those skilled in said culture or animal with an candidate antiviral agent, and
the art will know of other suitable carriers for binding determining the effect of said agent on said virus or its
monoclonal antibodies, or will be able to ascertain such infection of said culture or animal. An example of Such an
using routine experimentation. antiviral agent comprises an EMCR-CoV virus-neutralising
0057 The invention also provides a method for serologi antibody, or functional component thereof, as provided
cally diagnosing an EMCR-CoV virus infection of a mam herein, but antiviral agents of other nature are obtained as
mal comprising determining in a sample of said mammal the well.
presence of an antibody specifically directed against an 0063. The invention also provides use of an antiviral
EMCR-CoV virus or component thereof by reacting said agent according to the invention for the preparation of a
sample with a proteinaceous molecule or fragment thereofor pharmaceutical composition, in particular for the prepara
an antigen according to the invention tion of a pharmaceutical composition for the treatment of
0058 Methods and means provided herein are particu atypical pneumonia, especifically when caused by an
larly useful in a diagnostic kit for diagnosing an EMCR-CoV EMCR-CoV virus infection, and provides a pharmaceutical
virus infection, be it by Virological or serological diagnosis.
Such kits or assays may for example comprise a virus, a composition comprising an antiviral agent according to the
nucleic acid, a proteinaceous molecule or fragment thereof, invention, useful in a method for the treatment or prevention
an antigen and/or an antibody according to the invention. of an EMCR-CoV virus infection or atypical pneumonia,
0059 Use of a virus, a nucleic acid, a proteinaceous said method comprising providing an individual with Such a
molecule or fragment thereof, an antigen and/or an antibody pharmaceutical composition.
according to the invention is also provided for the produc 0064. The invention also comprises an animal model
tion of a pharmaceutical composition, for example for the usable for testing of prophylactic and/or therapeutic methods
treatment or prevention of EMCR-CoV virus infections and/or preparations. It is hypothesized that apes can be
and/or for the treatment or prevention of atypical pneumo infected with the EMCR-CoV virus, thereby showing clini
nia, in particular in humans. Preferably a peptide comprising cal symptoms, and more importantly, similar tissue mor
part of the amino acid sequence of the spike protein as phology as found in humans Suffering from atypical pneu
depicted in the relevant translations of FIG. 1, is used for the monia caused by the EMCR-CoV virus. Subjecting apes to
preparation of a therapeutic or prophylactic peptide. Also a prophylactic or therapeutic treatment either before or
preferably, a protein comprising the amino acid sequence of during infection with the virus will have a good and useful
the spike protein as depicted in the relevant translations of predictionary value for application of such a prophylaxis or
FIG. 1, is used for the preparation of a sub-unit vaccine. therapy in human Subjects.
Furthermore, the nucleocapsid of Coronaviruses, as depicted
in the translation of FIG. 1, is known to be particularly 0065. The invention is further explained in the Examples
useful for eliciting cell-mediated immunity against Coro without limiting it thereto.
naviruses and can be used for the preparation of a Sub-unit
vaccine. FIGURES LEGENDS
0060 Attenuation of the virus can be achieved by estab
lished methods developed for this purpose, including but not 0.066 FIG. 1: Nucleotide sequences from parts of the
limited to the use of related viruses of other species, serial EMCR-CoV virus. Also included are the putative amino acid
passages through laboratory animals or/and tissue/cell cul sequences of polypeptides.
tures, serial passages through cell cultures at temperatures 0067 FIG. 2: Phylogenetic relationship for the nucleotide
below 37° C. (cold-adaption), site directed mutagenesis of
molecular clones and exchange of genes or gene fragments sequences of isolate EMCR-CoV with its closest relatives
between related viruses. genetically. Phylogenetic trees were generated by maximum
0061 A pharmaceutical composition comprising a virus, likelihood analyses using 100 bootstraps and 3 jumbles. The
a nucleic acid, a proteinaceous molecule or fragment scale representing the number of nucleotide changes is
thereof, an antigen and/or an antibody according to the shown for each tree. FIG. 1a. Maximum likelihood tree of
invention can for example be used in a method for the matrix gene nucleotide sequences. Numbers in trees repre
treatment or prevention of an EMCR-CoV virus infection sent bootstrap values. The scale bar roughly reflects 10%
and/or a respiratory illness comprising providing an indi nucleotide differences between related sequences. FIG. 1b.
vidual with a pharmaceutical composition according to the Maximum likelihood tree of nucleocapsid gene nucleotide
invention. This is most useful when said individual com sequences. Numbers in trees represent bootstrap values. The
prises a human. Antibodies against EMCR-CoV virus pro scale bar roughly reflects 10% nucleotide differences
teins, especially against the spike protein of EMCR-CoV between related sequences.
virus, preferably against the amino acid sequence as 0068 FIG. 3: Similarity matrices indicating amino acid
depicted in the translation in FIG. 1, are also useful for identity for the putative Replicase 1a, Replicase 1b, Repli
prophylactic or therapeutic purposes, as passive vaccines. It case 1ab, Spike, Orf E. Matrix and Nucleocapsid proteins
is known from other coronaviruses that the spike protein is 3a-g, respectively), and for the putative Matrix protein and
a very strong antigen and that antibodies against Spike Nucleoprotein (3h and 3i resp.) between the EMCR-CoV
protein can be used in prophylactic and therapeutic vacci virus and closely related coronaviruses. See text for abbre
nation. viations.
0062. The invention also provides method to obtain an 0069 FIG. 4 Alignments with various coronaviruses: 5'
antiviral agent useful in the treatment of atypical pneumonia untranslated region genomic sequence (a): Putative orf 1 a
comprising establishing a cell culture or experimental ani amino acid sequence (b); Putative orf 1b amino acid
mal comprising a virus according to the invention, treating sequence (c); Putative orf 1ab amino acid sequence (d);
US 2008/0044426 A1 Feb. 21, 2008
Putative Spike amino acid sequence (e); Putative orf 4a Specimen Collection
amino acid sequence (f); Putative orf 4ab amino acid 0077. In order to find virus isolates nasopharyngeal aspi
sequence (g); Putative orf E amino acid sequence (h);
Putative Matrix amino acid sequence (i); Putative Nucle rates, throat and nasal Swabs, broncheo alveolar lavages,
oprotein amino acid sequence (); Putative 3' untranslated serum and plasma samples, and stools preferably from
genomic sequence (k); See text for abbreviations. mammals such as humans, carnivores (dogs, cats, mustellits,
Seals etc.), horses, ruminants (cattle, sheep, goats etc.), pigs,
rabbits, birds (poultry, ostriches, etc) should be examined.
EXAMPLES From birds cloaca Swabs and droppings can be examined as
well. Sera should be collected for immunological assays,
Specimen Collection such as ELISA, molecular-based assays, such as RT-PCR
and virus neutralisation assays. Collected virus specimens
0070 Virus was collected from an 8 month old patient may be diluted with 5 ml Dulbecco MEM medium (Bio
Suffering from pneumonia using nasal Swabs. Whittaker, Walkersville, Md.) and thoroughly mixed on a
Virus Isolation and Culture
Vortex mixer for one minute. The Suspension is thus centri
fuged for ten minutes at 840xg. The sediment is spread on
0071. Throat swabs were dipped into a culture of tMK a multispot slide (Nutacon, Leimuiden, The Netherlands) for
cells and passaged four times. Virus was then in Vero-118 immunofluorescence techniques, and the Supernatant is used
for virus isolation.
cells. One litre of virus containing cell culture Supernatant
was harvested, and the virus was pelleted in an ultracentri Virus Isolation
fuge and the virus pellet was resuspended 1 ml PES.
RNA Isolation
0078 For virus isolation Vero-118 cells or tMK cells
(RIVM, Bilthoven, The Netherlands) were cultured in 24
0072 RNA was isolated from the supernatant of infected well plates containing glass slides (CoStar, Cambridge, UK),
cell cultures or Sucrose gradient fractions using a High Pure with the medium described below supplemented with 10%
RNA. Isolation kit according to instructions from the manu fetal bovine serum (BioWhittaker, Vervier, Belgium).
facturer (Roche Diagnostics, Almere. The Netherlands). Before inoculation the plates were washed with PBS and
supplied with Eagle's MEM with Hanks' salt (ICN, Costa
Sequencing mesa, CA) supplemented with 0.52/liter gram NaHCO,
0.025 M Hepes (Biowhittaker), 2 mM L-glutamine (Bio
0073 Purified RNA was sent to BaseClear holding BV whittaker), 200 units/liter penicilline, 200 ug/liter strepto
(Leiden, The Netherlands) for sequencing. mycine (BioWhittaker), 1 gram/liter lactalbumine (Sigma
Aldrich, Zwijndrecht, The Netherlands), 2.0 gram/liter
Phylogenetic Analyses D-glucose (Merck, Amsterdam, The Netherlands), 10 gram/
liter peptone (Oxoid, Haarlem, The Netherlands) and 0.02%
0074. Nucleotide sequences were aligned using Clustal trypsine (Life Technologies, Bethesda, Md.). The plates
W running under BioEdit version 5.0.9. Maximum likeli were inoculated with Supernatant of the patient samples, 0.2
hood trees were created using the Seqboot and DNA-ML ml per well in triplicate, followed by centrifuging at 840xg
packages of Phylip 5.6 using 100 bootstraps and 3 jumbles. for one hour. After inoculation the plates were incubated at
The consensus trees were calculated using the Consense 37° C. for 1-7 days and cultures were checked daily for CPE.
package of phylip 5.6. These consensus trees were used as Extensive CPE was generally observed within 5-10 and
usertree in DNA-ML to recalculate the branch lengths from included detachment of cells from the monolayer.
the original sequences.
0075. The sequences of EMCR-CoV were compared Virus Culture
with those of reference viruses representing each species in (0079 Sub-confluent monolayers of tMK cells or Vero
the four groups of coronaviruses. These were: human coro clone 118 cells in media as described above were inoculated
navirus 229E (229E), af304460; porcine epidemic diarrhea with supernatants of samples that displayed CPE or with
virus (PEDV) af353511; transmissible gastroenteritis virus samples taken from a patient.
(TGEV), aj271965; bovine coronavirus (BoCoV),
af220295; murine hepatitis virus (MHV), af201929; avian RNA Isolation
infectious bronchitis virus (AIBV), m95169, Canine coro
navirus (CaCoV), d1309.6; feline coronavirus (FeCoV), 0080 RNA was isolated from the supernatant of infected
ay204704; porcine respiratory coronavirus (PRCoV), cell cultures or Sucrose gradient fractions using a High Pure
Z24675; human coronavirus OC43 (OC43), m76373, RNA. Isolation kit according to instructions from the manu
114643, m933990; porcine haemagglutinating encephalo facturer (Roche Diagnostics, Almere. The Netherlands).
myelitis virus (HEV), ay078417; rat coronavirus (RtCoV) RNA can also be isolated following other procedures known
af207551) References for the viruses are the numbers of the in the field (Current Protocols in Molecular Biology).
NCBI catalog (http://www.ncbi.nlm.nih.gov/entrezo).
Sequence Analysis
0076. In general, coronaviruses, such as EMCR-CoV can
be isolated and identified according to the following proto I0081 Sequence analyses were performed by BaseGlear
col: holding BV (Leiden, The Netherlands).
US 2008/0044426 A1 Feb. 21, 2008
SEQUENCE LISTING
The patent application contains a lengthy “Sequence Listing section. A copy of the “Sequence Listing is available in
electronic form from the USPTO web site (http://seqdata.uspto.gov/?pageRequest=docDetail&DocID=US20080044426A1).
An electronic copy of the “Sequence Listing will also be available from the USPTO upon request and payment of the
fee set forth in 37 CFR 1.19(b)(3).
1. An isolated essentially mammalian positive-sense 15. A method for serologically diagnosing an EMCR-CoV
single stranded RNA virus (EMCR-CoV) comprising the infection of a mammal comprising determining in a sample
sequence of FIG. 1 or homologues thereof. of said mammal the presence of an antibody specifically
2. An isolated positive-sense single stranded RNA virus directed against an EMCR-CoV virus or component thereof
(EMCR-CoV) belonging to the Coronaviruses and identifi by reacting said sample with a proteinaceous molecule or
able as phylogenetically corresponding thereto by determin fragment thereof according to claim 9.
ing a nucleic acid sequence of said virus and testing it in 16. A diagnostic kit for diagnosing an EMCR-CoV infec
phylogenetic tree analyses wherein maximum likelihood tion comprising a virus according to claim 1.
trees are generated using 100 bootstraps and 3 jumbles and 17. Use of a virus according to claim 1 for the production
finding it to be more closely phylogenetically corresponding of a pharmaceutical composition.
to a virus isolate having the sequences as depicted in FIG. 18. Use according to claim 17 for the production of a
1 than it is corresponding to a virus isolate of PEDV (porcine pharmaceutical composition for the treatment or prevention
epidemic diarrhea virus), HCoV-229E (human coronavirus of an EMCR-CoV virus infection.
229E), PRCoV (porcine respiratory coronavirus), TGEV 19. Use according to claim 17 for the production of a
(transmissible gastroenteritis virus), CaCoV (Canine coro pharmaceutical composition for the treatment or prevention
navirus) and FeCoV (feline coronavirus). of atypical pneumonia.
3. A virus according to claim 1 wherein said nucleic acid 20. A pharmaceutical composition comprising a virus
sequence comprises an open reading frame (ORF) encoding according to claim 1.
a viral protein of said virus. 21. A method for the treatment or prevention of an
4. A virus according to claim 3 wherein said open reading EMCR-CoV virus infection comprising providing an indi
frame is selected from the group of ORFs encoding the viral vidual with a pharmaceutical composition according to
replicase, nuclear capsid protein, matrix protein and the claim 20.
spike protein.
5. A virus according to claim 1 isolatable from a human 22. A method for the treatment or prevention of atypical
with atypical pneumonia. pneumonia comprising providing an individual with a phar
6. An isolated or recombinant nucleic acid or EMCR-CoV maceutical composition according to claim 20.
virus-specific functional fragment thereof obtainable from a 23. A viral replicase encoded by an RNA sequence
virus according to claim 1. comprising the indicated sequences, or homologues thereof
7. A vector comprising a nucleic acid according to claim as depicted in FIG. 1.
6. 24. A viral spike protein comprising the indicated amino
8. A host cell comprising a nucleic acid according to claim acid sequence as depicted in FIG. 1, or a homologue thereof.
6. 25. A viral nuclear capsid protein encoded by an RNA
9. An isolated or recombinant proteinaceous molecule or sequence comprising the indicated sequence as depicted in
EMCR-CoV virus-specific functional fragment thereof FIG. 1 or a homologue thereof.
encoded by a nucleic acid according to claim 6. 26. A viral nsp 3 or envelope protein encoded by an RNA
10. An antigen comprising a proteinaceous molecule or sequence comprising the indicated sequence as depicted in
EMCR-CoV virus-specific functional fragment thereof FIG. 1, or a homologue thereof.
according to claim 9. 27. A nucleic acid sequence which comprises one or more
11. An antibody specifically directed against an antigen of the sequences coding for separate viral proteins as
according to claim 10. depicted in FIG. 1 or a nucleic acid sequence which can
12. A method for identifying a viral isolate as an EMCR hybridise with any of these sequences under stringent con
CoV virus comprising reacting said viral isolate or a com ditions.
ponent thereof with an antibody according to claim 11. 28. A host cell comprising a vector according to claim 7.
13. A method for identifying a viral isolate as an EMCR 29. A method for virologically diagnosing an EMCR-CoV
CoV virus comprising reacting said viral isolate or a com infection of a mammal comprising determining in a sample
ponent thereof with a nucleic acid according to claim 6. of said mammal the presence of a viral isolate or component
14. A method for virologically diagnosing an EMCR-CoV thereof by reacting said sample with an antibody according
infection of a mammal comprising determining in a sample to claim 11.
of said mammal the presence of a viral isolate or component 30. A method for serologically diagnosing an EMCR-CoV
thereof by reacting said sample with a nucleic acid accord infection of a mammal comprising determining in a sample
ing to claim 6. of said mammal the presence of an antibody specifically
US 2008/0044426 A1 Feb. 21, 2008
directed against an EMCR-CoV virus or component thereof 39. Use of an antigen according to claim 10 for the
by reacting said sample with an antigen according to claim production of a pharmaceutical composition.
10.
31. A diagnostic kit for diagnosing an EMCR-CoV infec 40. Use of an antibody according to claim 11 for the
tion comprising a nucleic acid according to claim 6. production of a pharmaceutical composition.
32. A diagnostic kit for diagnosing an EMCR-CoV infec 41. A pharmaceutical composition comprising a nucleic
tion comprising a proteinaceous molecule or fragment acid according to claim 6.
thereof according to claim 9. 42. A pharmaceutical composition comprising a vector
33. A diagnostic kit for diagnosing an EMCR-CoV infec according to claim 7.
tion comprising an antigen according to claim 10. 43. A pharmaceutical composition comprising a host cell
34. A diagnostic kit for diagnosing an EMCR-CoV infec
tion comprising an antibody according to claim 11. according to claim 8.
35. Use of a nucleic acid according to claim 6 for the 44. A pharmaceutical composition comprising a proteina
production of a pharmaceutical composition. ceous molecule or fragment thereof according to claim 9.
36. Use of a vector according to claim 7 for the production 45. A pharmaceutical composition comprising an antigen
of a pharmaceutical composition. according to claim 10.
37. Use of a host cell according to claim 8 for the 46. A pharmaceutical composition comprising an anti
production of a pharmaceutical composition.
38. Use of a proteinaceous molecule or fragment thereof body according to claim 11.
according to claim 9 for the production of a pharmaceutical
composition.

US20080044426A1 (Novel Atypical Pneumonia-Causing Virus)

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(NL); Theodorus Marinus Publication Classification

mm. 5'UTR mummim

TTAAAAACTCTTAAGCT ACACTTACT CAATGGTTTTAGGTAATTGTGCCAAACGTAACGTTTTTGGTAAAATTCTIGATGT

GTTGAt ATAATTGAACAACCTTTTGGGGAAGTTGAACATGCGCTCT CAATTAGACAACCTTTTTCTTTTTCTTTTAGAGAIGAATTGGGTGT

AACGACTGTTTTAAGAATGATAATGTAGTTTTGAAAATTACTGAAGATGGTATTA ATGT TAAAGATGTfGTTGTTGAGTCTTCTAAGTCACT

CATAAAGGTGTTT GTTTTGGTTTTGATAAATGGTATGTTAATGATGGACGTGTTGATGACGGTTACATTT GTGGTGATGGTCTTATAGACCT

GT AGTGGGCTCATGTTGGTTCTGATTTT ACTGGTAGTGTTTATGGTAATTTTGATGACCAACC TAGTTTGCAAGTTGAGAGTGCCAACCTT

TCAAATTCAAACTGTTGATT CTTCT CAAGGTAGTGAGTATGATTATGTAATCTATGCACAAACTTCTGACACTGCACATGCTTGCAATGTAA

TGGTTGGGTATGGATGTTGAAAGTGCT CATGTTTGTGGCGATAACATAGGTACTAATGTTCCTTTACAGGTTGGTTTTT CAAATGGTGT TAA

TCT CAACTAAATGAAACTTTTCTTGATTTTGCTTATTTTGCCCCTGGTTTCTTGCTTTTCTACATGTAACAGTAATGCTAGTATT ICTATGT

GTAGTACT TCAAGACA ACAGTCTCGCACTCGTTCTGATTCTAACCAGTCTTCTTCAGAT CTTGTTGCTGCTGTTACTTTGGCTTTAAAGAAC

TTAGGTTTTGATA ACCAGTCGAAGT CACCAGTTCTTCTGGTACTCCACCCTAAGAAACCAATAAGCCTCTCTCAACCCAGGGCTGA

.. ..... . ........ . .. ..... . .... ...... .. ..... .. ..... .

... ..... . .... .. ... .... . .. ..... . .. ..... .. ..... .

... ..... . .... ..... . ... ...... .. ..... .. ......

... ..... . .... ..... .. .. . .. .. . .. .. ... .. . ... .. ....

......... . .... ..... . .. ..... . . . .. .. . . . .. . ... . .

.. ..... . .. .. .... . .... ... . . .... . .. .. . .. . . . .... . . . .. . . ... .

... .... . .. .. ..... . . ... .... . . ... ..... . .... .. . .. . . ... .. .. . .

.. . ..... ... ..... . ... ..... .

k. 5'untranslated region (genomic sequence)

229E5' UTR ACTTAAGTAC CTTATCTATC TACAGATAGA AAAGTTGCTT -TTTAGACTT TGTGTCTACT

.... ...... .... ......... .... . .... . .... . . .. . .... . .. . . .... .

... ...... .. .... . .. .....) .. ..... . .. ..... .

NOVELATYPCAL PNEUMONA-CAUSING each isolate may have a somewhat different percentage

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