DNA Replication

AP Biology 2007-2008
 1953 article in Nature
Watson and Crick

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 Double helix structure of DNA

“It has not escaped our notice that the specific pairing we have postulated
immediately suggests a possible copying mechanism for the genetic
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material.” Watson & Crick
 Directionality of DNA
 You need to PO4 nucleotide
number the
carbons!
 it matters! N base

5 CH2
This will be O
IMPORTANT!!
4 ribose 1

3 2
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OH
 5
The DNA backbone PO4
 Putting the DNA
backbone together base
5 CH2
 refer to the 3 and 5 O
4 1
ends of the DNA C
3 2
 the last trailing carbon O
–
O P O
Sounds trivial, but…
O base
this will be 5 CH
2
IMPORTANT!! O
4 1

3 2
OH
AP Biology 3
 Anti-parallel strands
 Nucleotides in DNA
backbone are bonded from
phosphate to sugar
5 3
between 3 & 5 carbons
 DNA molecule has
“direction”
 complementary strand runs

in opposite direction

AP Biology 3 5
 Bonding in DNA
hydrogen
bonds
5 3

covalent
phosphodiester
bonds

3
5

….strong or weak bonds?

How
AP do the bonds fit the mechanism for copying DNA?
Biology
 Base pairing in DNA
 Purines
 adenine (A)
 guanine (G)

 Pyrimidines
 thymine (T)
 cytosine (C)

 Pairing
 A:T
 2 bonds
 C:G
 3 bonds
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 Copying DNA
 Replication of DNA
 base pairing allows
each strand to serve
as a template for a
new strand
 new strand is 1/2

parent template &

1/2 new DNA

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 Let’s meet
the team…
DNA Replication
 Large team of enzymes coordinates replication

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 Replication: 1st step
 Unwind DNA
 helicase enzyme
 unwinds part of DNA helix
 stabilized by single-stranded binding proteins
helicase

single-stranded binding proteins

AP Biology replication fork
 Replication: 2nd step
 Build daughter DNA
strand
 add new
complementary bases
 DNA polymerase III

But…
Where’s the
We’re missing
ENERGY
DNA something!
for the bonding!
Polymerase III What?

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 Energy of Replication
Where does energy for bonding usually come from?
We come
with our own
energy!
You energy
remember energy
ATP!
Are there
otherenergy
other ways
to get energy
nucleotides?
out
You of
betit?
!

And we
leave behind a
CTP
TTP
GTP
ATP nucleotide! CMP
TMP
GMP
AMP
ADP
AP Biology modified nucleotide
 Energy of Replication
 The nucleotides arrive as nucleosides
 DNA bases with P–P–P
 P-P-P = energy for bonding
 DNA bases arrive with their own energy source
for bonding
 bonded by enzyme: DNA polymerase III

ATP GTP TTP CTP

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 5 3

Replication energy
DNA
 Adding bases Polymerase III
 can only add energy
nucleotides to DNA
3 end of a growing Polymerase III
DNA strand energy
DNA
 need a “starter”
Polymerase III
nucleotide to
bond to
DNA
energy
 strand only grows Polymerase III
53
B.Y.O. ENERGY!
The energy rules 3 5
the process
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5 3 5 need “primer” bases to add on to 3

energy
no energy


to bond
energy
energy

energy
energy

ligase
energy

energy

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3 5 3 5
 Okazaki

Leading & Lagging strands

Limits of DNA polymerase III
 can only build onto 3 end of
an existing DNA strand 5


f r ag ments
ki
Okaza 5
3 5 5 3
3
5 Lagging strand
3
ligase
growing 3
replication fork
5
Leading strand

Lagging strand

3 5

3
DNA polymerase III
 Okazaki fragments
 joined by ligase Leading strand
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 “spot welder” enzyme  continuous synthesis
 Replication fork / Replication bubble
3 5

5 3

DNA polymerase III

leading strand
5
3
3 5
5 5
5 3 3
lagging strand

3 5
5
3 lagging strand leading strand
5 growing
3 replication fork 5
5 growing
replication fork 5
leading strand 3
lagging strand
3
5
5 5

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 Starting DNA synthesis: RNA primers
Limits of DNA polymerase III
 can only build onto 3 end of
an existing DNA strand 5

3 5 3
5
3
3 5

growing 3 primase
replication fork DNA polymerase III
5

RNA 5

RNA primer 3

 built by primase
 serves as starter sequence

AP for DNA polymerase III

Biology
 Replacing RNA primers with DNA
DNA polymerase I
 removes sections of RNA DNA polymerase I
primer and replaces with 5

DNA nucleotides 3

3
5 ligase
growing 3
replication fork
5

RNA 5

3

But DNA polymerase I still

can only build onto 3 end of
an existing DNA strand
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 Houston, we
have a problem!
Chromosome erosion
All DNA polymerases can
only add to 3 end of an DNA polymerase I
existing DNA strand 5

3

3
5
growing 3
replication fork DNA polymerase III
5

RNA 5

Loss of bases at 5 ends 3

in every replication
 chromosomes get shorter with each replication
 limit
AP Biologyto number of cell divisions?
 Telomeres
Repeating, non-coding sequences at the end
of chromosomes = protective cap
5
 limit to ~50 cell divisions
3

3
5
growing 3 telomerase
replication fork
5

5
Telomerase
TTAAGGG TTAAGGGTTAAGGG
 enzyme extends telomeres 3

 can add DNA bases at 5 end

 different level of activity in different cells
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 high in stem cells & cancers -- Why?
 Replication fork
DNA
polymerase III lagging strand
DNA
polymerase I
3’
Okazaki primase
fragments 5’
5’ ligase
3’ 5’ SSB

3’ helicase

DNA
polymerase III
5’ leading strand
3’
direction of replication
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SSB = single-stranded binding proteins
 DNA polymerases
 DNA polymerase III
 1000 bases/second! Roger Kornberg
2006
 main DNA builder

 DNA polymerase I
 20 bases/second
 editing, repair & primer removal

DNA polymerase III Arthur Kornberg

enzyme 1959

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 Editing & proofreading DNA
 1000 bases/second =
lots of typos!
 DNA polymerase I
 proofreads & corrects typos
 repairs mismatched bases
 removes abnormal bases
 repairs damage
throughout life
 reduces error rate from
1 in 10,000 to
1 in 100 million bases

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 Fast & accurate!
 It takes E. coli <1 hour to copy
5 million base pairs in its single
chromosome
 divide to form 2 identical daughter cells
 Human cell copies its 6 billion bases &
divide into daughter cells in only few hours
 remarkably accurate
 only ~1 error per 100 million bases

 ~30 errors per cell cycle

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 What does it really look like?

3
4

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 Any Questions??

AP Biology 2007-2008
 For print version

AP Biology 2007-2008
 Energy of Replication
Where does energy for bonding usually come from?
We come
with our own
energy!
You
remember energy
ATP!
Are there
other ways
to get energy
out of it?

And we
leave behind a
ATP
TTP
GTP nucleotide! TMP
GMP
AMP
ADP
AP Biology modified nucleotide
 5 3

Replication energy
DNA
 Adding bases Polymerase III
 can only add
nucleotides to
3 end of the
growing DNA strand
 need a primer
nucleotide to
bond to
 strand grows 53
B.Y.O. ENERGY!
The energy rules 3 5
the process
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 5 3 5 3

no energy


to bond

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3 5 3 5
 5 3 5 3

ligase

energy

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3 5 3 5
 Houston, we
have a problem!
Chromosome erosion
DNA polymerases can
only add to 3 end of DNA polymerase I
an existing DNA 5

strand 3

3
5
growing 3
replication fork DNA polymerase III
5

5

Loss of bases at 5 ends 3

in every replication
 chromosomes get shorter with each replication
 limit
AP Biologyto number of cell divisions?
 Replication fork

3’
5’
5’
3’ 5’
3’

5’
3’
direction of replication
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