Course Title: Zoology-II

Course Code: ZOL-302

Credit Hr.: 4 (3-1)
Class: BS Zoology 2nd Morning

Lecture 6 DNA Replication

By: Dr Ummara Batool

AP Biology 2007-2008
 Contents
 Models proposed for DNA Replication
 Semi-Conservative Model
 Step involved in replication
 Unwinding
 RNA Primase

 Building daughter DNA strand

 Replacement of RNA primer by DNA

Polymerase I
 End of replication

AP Biology
 DNA Replication
 Purpose: cells need to make a copy of
DNA before dividing so each daughter
cell has a complete copy of genetic
information
 3 proposed Models of Replication

AP Biology
 Meselson and Stahl Experiment

https://youtu.be/frQbfdRtzBM
https://www.youtube.com/watch?v=JeoegQaF8ig
AP Biology
 Semi-Conservative Model
 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

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

in opposite direction
THIS WILL CAUSE A
PROBLEM FOR
REPLICATION
AP Biology 3 5
 DNA Replication
 Large team of enzymes coordinates replication

AP Biology
 Replication: 1st step
 Unwind DNA
 helicase enzyme
 unwinds part of DNA helix
 stabilized by single-stranded binding proteins
 PREVENTS DNA MOLECULE FROM CLOSING!
 DNA gyrase
 Enzyme that prevents tangling upstream from the replication
fork
helicase gyrase

single-stranded binding proteins

AP Biology replication fork
 Replication: 2nd step
 RNA Primase
 Adds small section of RNA (RNA primer) to the
3’ end of template DNA
 Why must this be done?
 DNA polymerase 3 (enzyme that builds new DNA
strand) can only add nucleotides to existing strands
of DNA

AP Biology
 Replication: 3rd step
 Build daughter DNA strand
 add new complementary
bases
 With the help of the enzyme

DNA polymerase III

DNA
Polymerase III
AP Biology
 Replication: 4th step
 Replacement of RNA primer by DNA
 Done by DNA polymerase I

AP Biology
 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
AP Biology
 “spot welder” enzyme  continuous synthesis
 DNA replication on the lagging strand
RNA primer is added
 built by primase
 serves as starter sequence for DNA polymerase III

HOWEVER short segments called Okazaki fragments

are made because it can only go in a 5 3 direction

5

3 5 3
5
3
3 5

growing 3 primase
replication fork DNA polymerase III
5

RNA 5

AP Biology 3
 Replacing RNA primers with DNA
NEXT 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

STRANDS ARE GLUED

TOGETHER BY DNA LIGASE
AP Biology
 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
AP Biology
SSB = single-stranded binding proteins
 Formation of Replication
Bubbles
Replication occurs in both directions
along the length of DNA and both
strands are replicated
simultaneously.
This replication process generates
"replication bubbles"

AP Biology
 Replication Bubbles

AP Biology
 The DNA Polymerase Complex
A number of different DNA polymerase
molecules engage in DNA replication. These
share three important properties: (1) chain
elongation, (2) Processivity, and (3)
proofreading.
Chain elongation accounts for the rate (in
nucleotides per second) at which
polymerization occurs.
Processivity is an expression of the number of
nucleotides added to the nascent chain
before the polymerase disengages from the
template.
The proofreading function identifies copying
errors and corrects them
AP Biology
Prokaryotic Eukaryotic DNA
polymerases


AP Biology
 DNA Topo isomerases
Relief of super coils is dome by Topo
isomerases
Two types:
Topoisomerases I : acts by making a transient
single cut in the backbone of the DNA,
enabling the strands to swivel around each
other to remove the build-up of twists
Topoisomerase II (DNA Gyrase) acts by
introducing double standed breaks enabling
one double-stranded DNA to pass through
another, thereby removing knots and
entanglements that can form within and
between DNA molecules.

AP Biology
 Proof reading and Editing

DNA polymerase III add

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

AP Biology
 Termination of replication

In prokaryotes:
DNA replication terminates when
replication forks reach specific
“termination sites”.
the two replication forks meet each
other on the opposite end of the
parental circular DNA .

AP Biology
 Termination of replication

This process is completed in about 30

minutes, a replication rate of 3 x 105
bp/min in prokaryotes
The entire mammalian genome
replicates in approximately 9 hours,
the average period required for
formation of a tetraploid genome from
a diploid genome in a replicating cell.

AP Biology
 Reconstitution of Chromatin Structure

chromatin structure must be re-formed

after replication. Newly replicated DNA
is rapidly assembled into nucleosomes,
and the preexisting and newly
assembled histone octamers are
randomly distributed to each arm of the
replication fork.

AP Biology
 Telomeres

In eukaryotic replication, following

removal of RNA Primer from the
5’end of lagging strand; a gap is left.
This gap exposes DNA strand to
attack of 5’ exonucleases.
This problem is overcome by
Telomerase.

AP Biology
 Telomeres

Repeating, non-coding sequences at

the end of chromosomes = protective
cap
AP Biology limit to ~50 cell divisions
 Mechanism of action of Telomerase

The enzyme synthesizes

(TTAGGG)n repeats on to the Telomere
sequences, using an internal RNA template.

AP Biology
 Summary of Replication
• Unwinding- forms replication fork
• Primase- Synthesizes RNA primer
• Continuous synthesis -Leading strand
• Discontinuous synthesis – Lagging strand
(Okazaki fragments)
• Synthesis 5’-3’ direction
• Primers removed, nick sealed
• Proof reading by DNA polymerases
AP Biology
 References
 https://www.youtube.com/watch?
v=dKubyIRiN84
 https://www.nature.com/articles/nature01407
 https://www.khanacademy.org/science/ap-
biology/gene-expression-and-
regulation/replication/a/molecular-
mechanism-of-dna-replication
 https://courses.lumenlearning.com/boundless
-biology/chapter/dna-replication/
 https://youtu.be/4jtmOZaIvS0

AP Biology