!

"#$
%&'()*#+),"
D. Renovaldi, M.Sc.(Biomed)
Faculty of Medicine & Health
Universitas Muhammadiyah Jakarta
 DNA Replication

Purpose: cells need to make a copy of

DNA before dividing so each daughter
cell has a complete copy of genetic
information.
 Base-Pairing Enables DNA Replication
• In the preceding chapter, we saw that each strand of the DNA double helix
contains a sequence of nucleotides that is exactly complementary to the
nucleotide sequence of its partner strand.
• Each strand can therefore act as a template, or mold, for the synthesis of a
new complementary strand
 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
 -./01/2$1/$!"# hydrogen
bonds
5¢ 3¢

covalent
phosphodiester
bonds

3¢
5¢
 !"#$34/567818$-721/8$95$%7:;1<951./$,=121/8$
v Site where replication begins
v Strands are separated to allow
replication machinery contact with the
DNA
v Characterized by many A-T base pairs
because easier to break 2 H-bonds that
3 H-bonds
 #/51>:9=9;;7;$85=9/08
• Nucleotides in DNA backbone are
bonded from phosphate to sugar
between 5¢ & 3¢ carbons
• DNA molecule has “direction”
• DNA is synthesized in the 5’-to-3’
direction
• complementary strand runs in
opposite direction
New DNA Synthesis Occurs at Replication Forks
 The Replication Fork Is Asymmetrical

• The 5’-to-3’ direction of the DNA polymerization

mechanism poses a problem at the replication
fork.
• The sugar– phosphate backbone of each strand
of a DNA double helix has a unique chemical
direction, or polarity, determined by the way
each sugar residue is linked to the next, and
that the two strands in the double helix run in
opposite orientations.
• As a consequence, at the replication fork, one
new DNA strand is being made on a template
that runs in one direction (3’ to 5’), whereas the
other new strand is being made on a template
that runs in the opposite direction (5’ to 3’)
 How is DNA Synthesized?

• Simple addition of nucleotides along one strand, as

expected
ü Called the leading strand
ü DNA polymerase reads 3’ ® 5’ along the leading
strand from the RNA primer
ü Synthesis proceeds 5’ à 3’ with respect to the new
daughter strand
• Remember how the nucleotides are added!!!!! 5’ à 3’
 How is DNA Synthesized?

• Other daughter strand is also synthesized 5’®3’ because

that is only way that DNA can be assembled.
• However the template is also being read 5’®3’
• Compensate for this by feeding the DNA strand through
the polymerase, and primers and make many short
segments that are later joined (ligated) together
• Called the lagging strand
• Those short fragments are called as Okazaki Fragments
!"#$%7:;1<951./$
Large team of enzymes coordinates replication
Replication
Steps
 -Unwind DNA-
• Helicase enzyme %7:;1<951./?$@85$857:
unwinds part of DNA helix
and 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
replication fork
RNA Primase
%7:;1<951./?$A/0$857:
• Adds small section of RNA to act as primers to the 3’ end of
template DNA
Why must this be done?
• DNA polymerase 3 (enzyme that builds new DNA strand) can
only add nucleotides to existing strands of DNA
 %7:;1<951./?$B=0$857:

§ Build daughter DNA

strand
u With the help of the enzyme
DNA polymerase III

DNA
Polymerase III
 %7:;1<951./?$C56 857:

• Replacement of RNA primer by DNA (Removal of Primers)

• Done by DNA polymerase I
• Other enzymes needed to excise (remove) the primers
ü Nuclease – removes the RNA primer nucleotide by
nucleotide
ü Repair polymerase – replaces RNA with DNA
ü DNA ligase – seals the sugar-phosphate backbone by
creating phosphodiester bond
ü Requires Mg2+ and ATP
Proteins at a Replication Fork Cooperate to Form a
Replication Machine
 Other Necessary Proteins

• Helicase opens double helix and helps it

uncoil
• Single-strand binding proteins (SSBP) keep
strands separated – large amount of this
protein required
• Sliding clamp
ü Subunit of polymerase
ü Helps polymerase slide along strand
• All are coordinated with one another to
produce the growing DNA strand (protein
machine)
 Mistakes during Replication
(DNA Repair)
• Base pairing rules must be
maintained
ü Mistake = genome mutation, may
have consequence on daughter cells
• If wrong nucleotide is included
ü DNA Polymerase uses its
proofreading ability to cleave the
phosphodiester bond of improper
nucleotide
ü Activity 3’ à 5’
ü And then adds correct nucleotide and
proceeds down the chain again in the
5’ à 3’ direction
%7:;1<951./$.D$!"#
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
SSB = single-stranded binding proteins
 How about telomere?

A telomere is the end of a chromosome. Telomeres are made of

repetitive sequences of non-coding DNA that protect the chromosome
from damage. Each time a cell divides, the telomeres become shorter.
Eventually, the telomeres become so short that the cell can no longer
divide.

Telomerase replicates the ends of eucaryotic

chromosomes
References
 +E#"F3
Do you have any questions?
Find me at :
de.renovaldi@umj.ac.id

Wanna watch an action movie?

Here!
https://www.youtube.com/watch?v=TN
KWgcFPHqw&ab_channel=yourgenome