You are on page 1of 5

Journal

OJ Shellfish

Research, Vol. 17.

REGULA TIO

(l.

1. 159-163.

1998.

OF MYTILUS GALLOPROVI
CIALIS GL YCOGE PHOSPHORYLASE BY
GLUCOSE A D GL COSE-6-PHOSPHATE
J A SERRA O, PILAR S AREZ ALO SO,
eo LOPEZ AND L. ose R GARCIA MARTIN*

F E el L
. L CA BL

Department of Biochemi try


Genetics and 11I1/1111110/0g)'
Faculty of cience
University of Vigo. Spain
"Department of Biochemistry and Molecular Biology
Faculty of Phanuacy
University of Santiago de Compostela, Spain
ABSTRACT
A different glucose and glucose--phosphare
inhibition 01' glycogen phosphorylase frorn Mvtilus rnantle relative lO those
01' vertebrates has been demonstrated.
Glucosc and glucose--phosphate
do not inhibir the a forrn (pho phorylated form), but they are
cornpetitive inhibitors 1'01' the substrate glucose-l-phosphate
in the b form (unphosphorylated forrn), with Ki values of 190 and 0.2-0.3
mM. respectively. The resulis cast doubt on the role 01' glucose a, a regulator and sugge t that gluco e-6-pho phate is a major regulator
01' glycogen phosphorylase activity in Myli/lIs
mantle tissue because 01' its special relalion. hip with gameiogenesis.
KEY WORD:

Myli/lIs

galloprovincialis.

glycogcn phosphorylase.

I TROD CTIO

regulation.

glucose,

glucose--phosphate

form) i activated by MP. LMP and high levels of inorganic


phosphate. and i inhibited by ATP, ADP, G6P and UDPG (Dombrdi 1981). Gluco e also how an inhibitory regulation synergisti !O inhibition exerted by purine derivative (Cori and Cori
1940. Ka vin ky el al. 1978, Carabaza 1992). The binding of G6P
10 the enzyrne cau e the inhibition of enzyrnatic activity through a
ncgative heterotropic effect on the AMP binding (Wang el al. 1970).
The concentration of circulating gluco e has long been con idered respon ible for the regulation of glucose homeo tasi by the
liver. Liver pho phorylase a, as the glucose receptor, i largely
re pon ible for the overall control of hepatic glycogen metaboli m
(Kasvinsky el al. 1978). Gluco e bind to glycogen pho phorylase,
rendering it a better sub trate for pho phoryla e pho phata e.
Then, glycogen phosphorylase is dephosphorylated and inactivated. Glucose is lile molecule in liver tis ue that trigger the
inactivation of glycogen pho phoryla e.
Alth ugh rnantle ti sue in Mytilus rnay be considered analogous to the liver in vertebrate ,a
a storage organ of glucidic
re. erves in the form of glycogen, its physiological function may
differ. o the aciivity and regulation of the glycogenolitic pathways in the mantle tissue of thi organi m doe not appear to serve
solely as a ource of gluco e lO be u ed intracellularly via the
glu litic route and the enzymatic
y tems involved seem lO
present a characteri tic function different from that of vertebrales .
Jt i. known that glycogen i the major energy fuel in the mantle
of bivalve mollu e (Zandee el al. 1980) and that an evident relationship between it degradation and gametogenic developmenl
exi 1 (Gabbott 1975. Bayne el al. 1982). During this developmenl
there i a clear coincidence between the increase of saturated fatty
acid and the decrease of the glycogen contained in the mantle
before pawning (Zandee et al. 1980). A key enzyme in the glycogenolytic pathways i the glycogen phosphorylase, present in the
Mytilus galloprovincialis mantle in IWO forms, a and b (San Juan
el al. 1991, an Juan el al. 1995). In this paper, we de cribe the
different propertie relative ro glycolitic metabolite ' regulation of
Mytilus mantle glycogen phosphoryla e, which is probably because of their differeru physiological role.

The glycogen pho phoryla e can be regulated by eo aleru


rnodifcation (pho phorylation-depho phorylation) and by allosteric rnodulation exerted al ihe molecular level by various phy iological factor acting through conformati nal change of the enzyrne. These factor can aCl as po itive and negative effector or
inhibitors (Fischer et al. 1971. Graves and Wang 1972. Bu. by and
Radda 1976. Kreb and Beavo 1979. Krebs 1981. Fi cher 19 3.
Madsen 1986).
Four separate ligand binding sites are localized in glycogen
phosphorylase of vertebral e . The active ite, which bind. the substrate . p or glucose-I-phosphate
(G 1P) inducing a a tive conformation R (Withers el al. 1982) and the nonredu ing terminal glucosyl unit of glycogen. as well as glucose and uridin-5'diphosphoglucose (UDPG). inducing an inactive conforrnauon T
(Sprang el al. 1982, Mad en el al. 1983). The nucleotide ite or
AMP allosteric site, consisiing 01' three different ub ite . namely
the pho phoryl group, the ribosyl residue and the purine ba ebinding loci. This site represent the primary binding site of allosteric activators: AMP and LMP as well as inhibitors:
TP. ADP
and gluco e-6-pho phate (G6P) (Johns n el al. 1979, Johns n el al.
1987). Furtherrnore, the ub. trates (i.e .. G 1P and Pi.
DPG).
nucleoside analogues,
r even dinucleotides are all capable of
. econdary binding to the nucleotide 1 cus. ATP. ADP. and G6P
exert iheir effe 1 through AMP expul ion (Dombrdi 1981. Morange el al. 1976. Eguchi el al. 1977). The nucleo ide (or inhibitor)
ite bind nucleosides fir 1 of all: i.e., adeno ine, ino ine. etc ..
purine ba e analogue (i.e., caffeine and theophyline and dinu leotides). furthermore. it al o repre ents the econdary binding . ite
of nucleotide (Ka vin ky el al. 1978. John on el al. 1979). And
the glycogen storage ite (Dombrdi 19 l. ewgard el al. 19 9,
Johnson el al. 1989). In vertebrate , the unpho phorylated form (b

Correspondence 10 F. San Juan errano. Departamento de Biologfa Fundamental. Facultad de Ciencia .. Uni er idad de Vigo. Apdo. 784. 36200
Vigo. Spain.

159

160

SA

MATERIALS

JUAN

D METHODS

Animals

Sea rnu sel were supplied by a purification plant in Villagarca


de Arosa (Galieia,
Spain). Mussel with helllength between
7 and 10 cm were elected for the experiment . They were transponed lO the laboratory within 1 hr of collection. Their rnantle
were separated and lo red at -30C until u e.
Reagents

The ubstrates, enzyme. and eoenzymes were obtained from


Sigrna Chernical Company (St. Louis. MO) and Boehringer Mannheim (Mannhei m , Gerrnany).
Mag ne s iu rn acet ate , t r ishydroxyrnethyl aminornethane, irnidazole and other ehemieals and
salts were purehased from Merk (Darrnstadt, FRG).
Preparation

o/ Phosphorylase

Extracts

Phosphorylase b extraer was prepared frorn rnantle tissue


hornogenized in 2 volumes (w/v) of ice eold 40 mM tris-acetate
buffer pH 7.0, eontaining EOTA, imidazole and 2-mereaptoethanol in a concentration of 5 mM and 0.12 M KCI. For preparation of pho phorylase a extraer, a hornogenate in the same eonditions de eribed above. but without EOTA, wa incubated with 2
mM ATP, MgCI2 and CaCI2. In thi medium. endogenou phophoryla e kina e tran forms the pho phoryla e b lO its a form ( an
Juan el al. 1995). Both homogenate . coruaining b and a phosphoryla e forms respectively. were centrifuged at 150400 x ga, for 20
rnin and the pellet was di earded. The upernatant were dialyzed
again l 30 vol of extraction buffer containing 10 mM aF and
without KCI.
EIIlJ!me Determinations

Glyeogen pho phorylase aetivity wa mea ured spectrophotornetrically in the direetion of glyeogen breakdown, a de eribed in
San Juan el al. (1991). The reaction mixture for phosphoryla e b
contained 40 mM tris-acetate buffer pH 7.0, 5 rnM irnidazole, 2
mM EOTA, lA mM 2-ereaptoelhanol. 5 mM acetate-Mg, 5 ~M
glueose-I-6-diphosphate,
0.6 mM
AOP, 80 mM pho phate
buffer, pH 7.0, 1.6 mM AMP, 16 mg/mL glycogen, 0.5 I.U. of
glueo e-6-phosphate dehydrogena e. 0.1 J.U. of pho phoglucornula e. and enzyme preparation in a total volume of 1.0 mL. The
reaction mixture for pho phoryla. e a lacked EOTA and AMP. and
the glycogen and pho phate buffer concentrations were
mglmL
and 30 mM, re pectively.
Pho phoryla e activity wa as ayed in the direction of glycogen
ynthesi by mea uring the relea e of P, from G 1P. The reaetion
rnixture for the pho phoryla e b eontained 40 mM tri -a etate
buffer, pH 7.0.2 mM EOT A, .2 mg/mL glycogen, 1.6 mM AMP,
00 mM G-I-P, and enzyme preparation in a total volume of I mL.
The reaetion mixture for pho phoryla e a laeked EOTA and AMP.
and the G 1P eoneentration wa 50 mM. After incubation al 30C
for 45 mino the reaction was topped by the addition of 0.1 mL of
20% (w/v) SOSo After centrifuging, P, in the upernatant was
determined by the method of Saheki et al. (1985).
Dala Analysis

The inhibitory effect eau ed by everal concentration


of gluco e and G6P wa determined by a rnean of provoked modifications in the kinetic constants of glyeogen pho phoryla e in equilibrium. in relation to its ub trates. The inhibitor aetion meeha-

ERRANO

ET

L.

ni m wa deterrnined u ing Lineweaver-Burk double reeiprocal


and Hill plots. The inhibition constants were calculated from the
representation of double reeiproeal (-I/Kmap = -l/Km (1 + [IV
K,). When the pre. enee of an inhibitor di places the repre en lations 01' double reeiproeal from linearity, the repre entation of IIV
a oppo ed lO 1/[ )2 was used. In this rnanner, the illu tration is
linearized and the inhibition con tant may be calculated in these
ea. es, a. in tho: e de eribed above (Segel 1993).
Kinetic pararneters (Km or Scu and VmuX>were ealculated u ing
thc "Graf'it program (Erithacu Software Ltd .. Sraines, U.K.).
Result are given as mean EM. Eaeh date corre pond to rneans
of 4-5 determination
and regre sion coefficient of LineweaverBurk, and Hill plOIS were 0.96-0.99.
RESUL TS AND DI CUSSION

Because 01' the method used lO rneasure pho phorylase activity


in the direction of glycogen brcakdown, only the effect produced
by the gluco e and the UOPG eould be examined. Neither of the
lWO rnolecules present inhibitory effects on the enzyme (Iorrns a
and b) of Mytilus in this sense.
Glyeogen pho phoryla e (a-lA glyeogen, ortho-pho phate glueosyltransfera e E.e. 204.1.1) eatalyze the degradation of glyeogen lo glucose-t-phosphate.
Thi reaction i rever ible. but in the
cell the ynthe i of G 1P trongly favor the high Pi concentration . In vitro, the ratio [Pi]/[G 1P)that determined the equilibrium
con tant can be rnodified (i.e., inereasing the G 1P al saturating
concentration, 800 mM), di plaeing the reaetion in reverse direeiion. BUl the enzyme affinity for the ubstrates and effector and
the interactions between them are the ame. and they can be studicd in both direcLion .
In the rever e reaetion direction ( ynthesis of glyeogen), UOPG
neither showed any inhibitory effect, wherea the glueo e and G6P
present an evideru inhibition of form b relative to the G IP ubtrate, but they do n 1 affe t the activity of form a, whieh G6P even
seem lO actvate mildly (Fig . l. 2). Kinetic parameters of this
enzyme were determined using aliquots of the purified enzyme
(esscntially a San Juan el al. 1991) and u ing crude li ue extract,
before purification, giving identical result . Although glyeogen
pho phoryla. e reaction wa not rneasured in physiological direction, the re ults of this srudy show the Mytilus enzyrne affinity for
the e inhibiton and their effects on the conformational changes of
moleeular tructure, which affect to catalitic ite and its affinity for
all ub trate.
In relation t form b of the Mytilus enzyme, both moleeule
(glucose and G6P) follow a cornpetitive type pattern, only affecting the affinity of the enzyme by the G 1P ub trate. G6P increa es
the Km in an order of magnitude from 180 mM to 1.8 M when il
increa e conceniration from O lO 1.5 mM. The Ki obtained for lhi
inhibitor i between 0.2-0.3 mM (Fig. 3). a value similar to that
obtained in the adductor mu eJe of Pecten (Hata et al. 1987). Al o,
the glueo e produce a far lower increa e of Km for the G IP (up
to 217 mM) when it
n entration range from O to 20 mM,
alculating a Ki of 190 mM (Fig. 4). Both in the presenee of G6P
and of gluco e, an increa e in the Hill Index for the G I P frorn
values approaehing 1 10 value of 1.5 and 1.9. re pectively i
ob erved (Figs. 3. 4 insets).
The behavior of both forrn of enzyme in the mantle tis ue of
Mytitus in relation LO the e two inhibitor differ notably from
what is de cribed in the bibliography, particularly a. regard form
a and inhibition by glueose of forrn b.

REGULATIO

OF MYTILU

GLY

EN PHOSPHORYLA

161

lO,

,.1

.-.. ..

',t

.'

,..

,.1
t

00

10

20
'09 (GIP)""

"

'0

e
1

lo

().I-----..----+__

r).o

o. -

==~-L~)~~~

--l-

n'~

(l.")

(b~0~)~~~~--~(~I~I!.=4==~=~=)=0
1 . ()

r\
Figure 1. Double recprocal plot of initial velocity of pho phorylase a
as funclion of G lP concentration
in the pre ence of several gluco e
concentratons: ( ) O rnM, (e) 10 mM, (6) 20 111M.

G6P i. de. cribed as a typical allo teric inhibitor of form b of


the glycogen phosphoryla e in relati n 10 the AMP with which it
cornpetes for the nucleotide site (Mad en and hecko ky 1967) by
binding 10 the phosphate ubsite (Lorek el al. 19 4). Thi would
explain our results, becau e a le er bind of the AMP lO the nucleotide place would not actvate the bind of the substrate 10 the acti e
ccnter 10 the ame extent as if an inhibir r were la king, thus
causing an evident decrease in affinity. In turn, a. the bind of the

1~' l rn

Figure 30 Double recproca] plotsof initial velocity of phosphorylase b


as function of G I P concenlralion
in the presence of everal G6P concentrations:
( ) O 111M, (e) 0.5 mM. (6) 1.5 mMo Inset: HiII plot,

G 1P with the active center i le strengthened. thi ubstrate may


bind. in a econdary formo 10 the nucleotide place by [he subsite for
the P, (Dornbrdi
19 1), a indicated by the increase in the Hill
Index (Table 1)0
Gluco e i a well kn wn inhibitor of both form of the glycogen pho phoryla e, tabilizing the inactive T conformation by bind
10 the acti e center and redering it a better sub trate for pho phorylase pho phata e (Her 1976, Sprang el al. 1982, Mad en el al.
o

40

"

o.

>-o~

')0

40
t:

t
-)

"

-n
00

zc

tO

lO

Ioq{CIPJmlot

0+

_~-n

.:n..

~, ~

II

1 -

)~-

00

o.r
,1 F

O. '

1.0

r'IM

Figure 2. Double reciprocal plots of initial velocty of phosphorylasc


a
as function of G I P concentration
in the presence of several G6P concentrations:
( ) O mM, (e) 005 111M, (6) 1.5 111M.

o. 7
'/[()

::! 08

1 .

IP]

11

1<,2

Figure 4. Double reciprocatplotsof initial velocityof phosphorylase b


as funclion of G IP concentration
in the pre ence of several gluco e
coucentrations:
( ) O rnM, (e) 10 111M, (6) 20 111M. lnset: HiLl plot.

162

SAN
TABLE

AN SERRANO

1.

ET AL.

phosphorylase

Modification of kinetic parameters


of glycogen phosphorylase
(a and
b) from Mylilus mantle with respect to substrate G 1P in the
presence of several G6P and glucosc concentrations.

the sarne order.


physiological

nH

Vm (1 /mL)

whereas

direction

Taking
Km (111M)

for both, substrate (G 1P) and inhibitor

The value of both constants,

(glucose).

Km and Ki respectively,

the value of Km

is 95-fold

are within

for the glycogen

in

srnaller (San Juan et al. 1993).

into account the seasonal coruent of free tissue glucose

regulated regardless 01' the mobilization of glyco01' glucose--phosphate phosphaiase activity in the rnantle tissue of Mytilus, which involves an irnportant
difficulty for the gluco e to be the final expression 01' the glycogenolitic effector system (Cres po 1989). and the high values 01' Ki
(70-130

mM).

gen. the non-existence

b FORM
[glycogen]: 3.2 I11g/I11L
IAMP): 1.6 111M

IG6Pl
0.0 mM
0.5 mM
1.5 111M

0.475 3 x 10--'

182.38 2 x 10-2
296.59 6 x 10-'
1699.0 6 x 10-'

0.415 7 x 10
0.491 5 x 10--'

0.8
1.5
1.4

191.98 6 x 10-'
217.546x
10-'

0.445 3 x 10
0.395 5 x 10

1.3
1.9

[glucose]

10.0 111M
20.0 mM
a FORM
1: 3.2 rng/ml,

for this metabolite

obtained

the regulating role 01' the glucose


paihway lacks the meaning and importance
de scribed in vertebrate where il is the rnost important physiological inhibitor of phosphorylase (Glin rnann el al. 1970, Stalrnans
and Gevers 1981. Carabaza el al. 1992) as occurs in other invertebrate (in ects) where the regulatory role is in doubt (Applebaurn
in the glycogenolitic

1973. Van Marrewijk

[glycogen

Furthermore,

[G6PI
0.0 mM
0.5 mM
1.5 111M

1.1

0.123 4 x 100.157 4 x 10--'

1.49 5 x 10
0.89 1 x 10-'
0.81 4 x 10--'

0.1433x

0.6
0.6

10-'

1.127xI0--'
1.34 1 x 10-'

el al. 1988).

the most rnarked

with the regulating

main interaction

0.9
0.8

of G6P, it

effect

found in the mantle

1990), besides the high phosphoglucormnase


and

akada 1968) and the non-existence

phosphate pho sphatase

0.112 4 x 10--'

inhibitory

site of the enzyme and its low

Ki. inside the range of concernrations


(Ibarguren
(Bennett

[glucose]
10.0 mM
20.0 mM

in this work (190 111M), we rnay con-

sider that in the Mytilus mantle,

(Crespo

tissue

activity

01' glucose-6-

1989). lead us to attribute

a regu-

Results are expressed as mM (Km - SIl~) and IU/ml SEM (standard error

lating effect on this pathway to this rnolecule, in this lis ue, which
is more important than that of gluco e. as also eern lO occur in
the Iar-body 01' in ects (Applebaum
1973) and in yeasts (Sagardia

means).

el al. 1971).

1983, Carabaza et al. 1992). In ihe case 01' the Mytilus enzyme, we

observed

0.139

1 x 10-'

lO recently obtained but, unpublished data, we have


a higher proporrion of phosphorylase b in the rnantle
ti sue of Mytilus which i significantly
correlated with garnetogeAccording

have observed the inhibition


Iorrn a. The inhibition
is clearly a cornpetitive
active

on forrn b and the noninhibition

on

exerted by rhis rnolecule on the G 1P bound

type, both binding 10 the same area in the

center and nOI via negative

heterotropic

behave .. As before, the G 1P displaced

effects

by the inhibitor

active center tends to interact with the nucleotide

as G6P
frorn the

site, and consis-

tenily an increase in the Hill Index is observed (Table 1).


A curious fact, consiantly observed in the re ults obtained in
lhi

paper. is the greater affinity

of the form phosphorylated

G 1P ubstrate (the Km decrea e between


the sarne lime as it
of the affinity
forrn (Table
molecules

mximum

velocity

decrease four-fold

and mximum

velocity

of the non-pho

1). Moreover,

is particularly

becau e the inhibiiory

on the affinity

by the

122 and 2098 times), al


in rerrns

phorylaied

effect of b th

by the G 1P and since this

ni

development,

al limes when the concentration

glycogen

degradation

rnay be derivecl toward

enzymatic
energy

form is described

dernand,

pr bably needs lo Iransfer the glucidic


re erves in the forrnation

in short periods 01' time.

the activity nor the affinity

upports the hypothe i


ite than
cerner in the pho phorylated forrn of the Mytilus

of a greater interaction
with the active

enzyrne

of form a, this

of the G 1P with the nucleotide

(San Juan 1991).

The inhibitory
only,

do nOI affect either

effect

can be explained

irnportance

in reaction

by the low affinity

rever. e direction

of Mytilus glycogen

REFERE
Applebaurn, S. W. & H. M. Schlesinger. 1973. Regulan
body pho sphorylase. Biochem. 1. 135:37-41.
Bayne. B. L., A. Bubel, P. A. Gabbou, D. R. Livingston,

n of Locust falD. M. Lowe &

prolonged

inhibitor,

in vertebrates,
coruribution

the

by the substrate

can

condition

of glycogen degradation by ihese me-

regulation

paper. in comparison

may be considered
physiological

function

01' phosphorylitic

cogen in the gametogenic

10 lipidic

regard to this

of high energy dernand as stress

as shown in thi

of

when it very

corroborates

a with

which together with its high affinity

The different
tabolites,

of form

of low

period

re erve depo it

01' garnete . Also, thi

of the independence

10 a different

of glucose

in situations

during

time (Johnson el al. 1989), as it is the garnetogenesis.

duced. Apart from the fact that these inhibitors

ignificantly

as fundamental

bUI is maintained

respond al situations

is

the pentose phos-

phate pathway lO form nucleic acids and reductor power required


in ihe biosynthesis of lipids, during garnetogenic develcpment.
This SUppOl1S and justifie
the low affinity of forrn b by the
glucose and its special regulation by G6P. The activity of this

low, al o in the absence of inhibitors (Km


= 180 mlvl), the
arce phy i logical importan e of the glycogen
pho phorylase reaction of Mytilus in this direction rnay be de-

affinity

of G6P is al a

mnimum and G6P dehydrogena e activity i at a rnaximum. The e


data suggest that, al particular times, the phosphorolitic pathway 01'

with that described

as an adaptarional consequence
and provides new data on the

pathway

in the mobilization

process of bivalve

of gly-

molluscs.

CES
M. . Moore. 1982. Glycogen utili ation and garnetogenesis in Mytilus
edil lis L. Mar. 8iol. Lett. 3: 9-105.
Benneu, R. & H. 1. akada. 1968. Comparative carbohydrate rnetabolism

REG

LATIO

OF MYT/LUS

01' marine rnollusc. 1. The intermediury metabolivm 01' My/i1I1.\ californianus and Haliotis rufescens.
omp. Biocliem. Phvsiol. 24:787-797.
Busby. S. J. W. & G. R. Radda. 1976. Regulation 01' the glycogen phosphorylase systern from physical rneasurerncms 10 biological speculations. Curro Top. Cell Regul. 10:89-160.
Caraba/a.
.. C. J. Ciudad. S. Baque & J. J. Guinovart. 1992. Glucose ha,
10 be phosphorylated
10 actvate
glycogen syntha: e. but not 10 inuciivate glycogen phosphorylase in hepatocytes. Feb.\ Let. 296:21 1-214.
Cori. G. T. & C. F. Cori. 1940. The kinetics of the enzyrnatic synthesis 01'
glycogen from glucose-I-phosphate.
J. Biol. Chem. 135:733-742.
Crespo. C. A. 1989. Histofisiologfa de 1m, reservas bioenergticas en el
manto de Mvtitus galloprovincialis
Lmk. Doctoral Thesis, University
01' Santiago de Cornpo lela. Spain.
Dombrdi. V. 1981. Structural aspects 01' the caralytic and regulatory function 01' glycogen phosphoryla. e. 1/1/. J. Biochem. 13: 125-139.
Eguchi. c.. K. Suzuki & K. lrnahocguchi. 1977. Synthesis of AMP analogs
and their use for siudies on the allosteric site 01' rabbit rnuscle glycogen
phosphorylase. J. Biol. Cliem. 81: 140 1-141 1.
Fischer, E. H., L. M. G. Heilmeyer & R. H. Haschke. 1971. Phosphorylase
and the control 01' glycogen degradarion. Curr. Top. Cell Regul. 4:21
249.
Fischer. E. H. 1983. Cellular regularion by protein phosphorylaiion. BIIII.
II/.I/i/. Pasteur 81 :7-31.
Gabbott. P. A. 1975. Storage cycles in marine bivalve rnolluscs: a hypoihesis concerning the relationships between glycogen metabolism and
gametogenesis.
pp. 191-211./1/: H. Barnes (ed.).
inth European Marine Biology Symposium. Aberdeen
niv. Press.
Glinsrnann. W .. G. Pauk & E. Hern, 1970. Glycogen phosphorylase regulation in liver. Biochem. Biophvs. Res. Commun. 39:77-1--782.
Graves D. J.
J. H. Wang. 1972. o-glucun phosphorylases: chernical and
physical basi of cataly is and regulation. Enzyme 7:435-452.
Hata K.. Y. Yokoyarna. M. uda. M. Hata & K. Matsuda. 19 7. Purification and properties
of glycogen phosphorylase
Irorn the adductor
mu sclc of the callop. Patinopecten vessoensis. Comp. Biochem. Phvsiol. 87B:747-753.
Hers. H. G. 1976. The control 01' glycogen metabolisrn in the liver. AI/I/.
Re\'. Biochem. 45: 167-1 9.
lbarguren, 1. 1990. Control de la glucolisis en el mejilln gallego (My/ill/.I'
galloprovincialis
Lrnk). Doctoral Thesis,
niversity of antiago de
ornpostela. Spain.
Johnson. L. .. E. A. Stura, K. S. Wilson. M. S. P. Sansorn & 1. T. Weber.
1979. Nuclcotide binding 10 glycogen phosphorylase b in the cristal.v.
Mal. Biol. 134:639-653.
Johnson. L. .. K. R. Acharya, D. 1. Stuart, D. Bardford.
. G. Oikonomakos. J. Hajdu & K. M. Varvill, 1987. Phosphate-recognition
sites in
catalysis and control of glycogen phosphorylase b. Biochem. Soc.
Trans. 15:1001-1005.
Johnson, L. .. J. Hadju. K. R. Acharya, D. 1. tuart, P. 1. Mclaughlin.
. G. Oikonomakos & D. Bardford. 1989. Glycogen phosphorylase b.
In: G. Herve (ed.). Allo teric enzymes, CR
Press. Boca Ratn.
Florida.
Kasvinsky, P.1..
. B. Madsen. J. ygush & R. J. Fleuerick. 1978. The
regulalion of glycogen pho phoryla e a by nucleotide derivatives. Kinetic and X-ray cry tallographic
tudies, J. Biol. Chem. 253:33433351.
Krebs, E. G. & J. A. Beavo. 1979. Phosphorylarion-dephosphorylaricn
01'
enzyrne . AI/I/. Rev. Biochem. 4 :923-960.
Krebs. E. G. 1981. Pho phorylation and dephosphorylation
of gly ogen
pho phorylase: a prototype for reversible covaleru m dification, Curr.
Top. Cell. Regl/I. 18:401-419.
Lorek, A .. K. S. Wilson. M. S. P. anson. D.1. tuan, E. A. tura. J.
Jenkins. J. Hajdu & L. . J hn on. 1984. Allosteric intera tion of
glycogen pho phorylase b. Biochem. J. 218:45-60.

GLY

OGE

PHO PHORYLA E

163

Madscn.
. B. & . hechosky, 1967. llosteric properties 01' phosphor) lave b. 11.Comparison with a kinetic model. J. Biol. Chem. 242:33013307.
Mud-en.
. B...
hechosky & R. J. Fleuerick. 1983. Site-site interacti ns
in glycogen phosphorylave b EC-2.4.I.1 probed b) ligands specific for
cach site. Biochem. 22:4460-1465.
tadsen.
. B. 1986. Glycogen phosphory lase. pp. 365-393. 11/: P. O.
Boycr and E. G. Krebs (eds.). The enzyrnes: enzyme control by phosphorylation. Acadernic Press. e" York,
Morange. M .. F. Garca Blanco. B. Vanderbunder & H. Boc. 1976. AMP
analogucs: their funclion in the activation 01'glycogen phosphorylase b.
El/l'. J. Biochem. 65:553-563.
ewgard,
. B.. P. K. Hwang
R. J. Fletterick. 1989. The family 01'
glycogen phosphorylases:
struciure and funcuon. Crit. Rev. Biochem.
Mol. Biol. 24:69-99.
Sugardia, F.. 1. Gotay & M. Rodrtguez. 1971. Control properties 01' yeast
glycogen phosphorylase. Biochem. Biophvs. Res. COI1lI/l. 42:829-835.
Saheki. S .. A. Takeda & T. Shirna/u.
1985. Assay of inorganic phosphate
in thc mild pH range. suitable for rneusurernent 01' glycogen phosphoryluse activity, Al/al. Biochem. 148:277-281.
San Juan Serrano. F. 1991. aracterizacin
rnolecular y cintica de la
enzima glucgcno fosforila a (E. . 2.4.1.1) de manto de Mvtilus galloprovincialis
Lrnk. DOCl ral Thesis.
niver iry 01' Santiago de Composiela. pain.
an Juan errano, F.. M. Fernndcz Gonzlez. J. L. Snchez Lpez & L. O.
Gnrca Martin. 1991. Purification and rnolecular properties 01'glycogen
pho phorylase b frorn rnarule ti sue 01' mussel Mvtilus galloprovincialis. Comp. Biochem. Phvsiol, 98B:33-39.
San Juan Serrano. F.. M. Fernndez Gonzlez. J. L. nchez Lpez, &
L. O. Garca Martn. 1993. A kinetic -rudy on glycogen phosphorylase
b trorn the marule of Mvtilus gal/oprOlirrcialis.
Lrnk. Comp. Biochem.
Pliysiol. I06B:925-932.
un Juan errano, F.. M. Femndez Gon/lez. J. L. Snchez Lpe7 & L. O.
Garcia Martin. 1995. Molecular mechanisrn of the control 01' glycogenolysiv by calcium ion, and cyclic AMP in the mantle 01' Mytilus
gulloprovincialis
Lmk. Comp. Biochem. Physiol. IIOB:577-582.
cgcl, 1. H. 1993. Behavior and analysis of rapid equilibrium and steady<tate enzyrne systerns. 11/: Wiley-Inrerscience USA (ed.). Enzyme Kinctics.
Sprang, S. R.. E. J. Goldsrnith. R. J. Fleuerick. S. G. Withers & N. B.
Madsen. 1982. Catalytic site 01' glycogen phosphoryla e: structurc 01'
ihe T state and pecificity tor o-n-glucosc. Biochem. 21 :5364-5371.
talman . W. & G. Gevers. 1981. Comparative
in thc liver. Biochem. 1. 200:327-336.
Van

siudies of phosphorylase

Marrewijk.
W. J. A .. A. Th. 111. Van Den Broek & A. M. Th.
Beenakakers. 1988. lsolation and partial characterization 01' three form
of glycogen phosphorylase frorn fat body of Locusta migratorio. 11/ ect
Biochem. 18:37-l4.

Wang. J. H.. M. L. Shonka & D. J. Graves, 1965. Intluence of carbohydrates on phosphorylase structure and activity. 1. Activation by preincubation with glycogen. Biochem. 4:2296-2301.
Wang. J. H.. J. 1. Tu & F. M. Lo. 1970. Effect 01' glucose--phosphate
on
the nucleotide ite of glycogen phosphorylase b. A general approach for
negative hcterotropic interactions. J. Biol. Chem. 245:3115-3121.
Withers,
. G.. . B. Madsen & B. D. ykes. 1982. Covalently activated
glycogen phosphorylase: a phosphorus-Jt
NMR and ultracentrifugation analysis. Biochem. 21 :6716-6722.
Zandee, D. 1.. J. H. Kluytrnans, W. Zurburg
H. Pieters, 1980. Seasonal
variations in biochernical cornpo ition 01' Mvtilus edil lis with reference
10 energy rnetaboli m and gametogenesis.
Neth. 1. ea Res. 14: 1-29.