DNA REPLICATION

DNA: genetic material

• transmits biological information from parent to daughter cell,
from generation to generation

• stored information when expressed, result to a particular trait.

• central dogma of molecular biology

3 general processes :
 DNA replication has two requirements that
must be met:
1. DNA template

2. Free 3' -OH group

THREE STEPS IN DNA REPLICATION
• Initiation
• Elongation
• Termination
DNA Vocabulary
• DNA Gyrase (topoisomerase) – enzyme that untwists the double helix

• DNA B (helicase) – proteins that bind to the double stranded DNA

and stimulate the separation of two strands by
breaking the hydrogen bonds that hold the
nucleotides
• Single-stranded binding proteins – proteins that bind to the DNA as a
tetramer and stabilize the single
stranded structure that is
generated by the action of the
helicases. Replication is 100 times
faster when these proteins are
attached to the single-stranded
DNA
• Dna Polymerase I (Pol I)
- 3’ to 5’ exonuclease (proof-reading activity)
- 5’ to 3’ exonuclease (repair activity)

• Dna Polymerase III (Pol III)

- the enzyme that performs the 5’ to 3’ polymerase function
(elongation)

• Primase – synthesizes RNA primers

• DNA ligase – seals any gaps that remain between bases of the “new” DNA
by forming phosphodiester bonds between 3’ – hydroxyl and 5’
phosphate groups.
• Ort C (origin of replication) – a specific spot in the DNA molecule
where replication begins

• Initiator proteins – proteins that recruit molecules to pry apart the

two strands of DNA
DNA Replication in Prokaryotes
INITIATION
Step 1: DNA binds around initiator
proteins, then DNA B
(helicase) unwinds ort C
(origin of replication) and
extends the single
stranded region for
copying.

Step 2: Single strand binding

proteins (SSB) binds to the
single stranded region to
protect it from breakage
and to prevent it from
renaturing.
Step 3: DNA gyrase
(topoisomerase)
untwists the DNA
ahead of the replication
fork

Step 4: The enzyme DNA

primase then attaches
to the DNA and
synthesizes a short RNA
primer to initiate
synthesis of the leading
strand of the
replication fork
DNA polymerase III
ELONGATION
Step 5: DNA
polymerase III
extends the RNA
primer made by
primase. All DNA
polymerases make
DNA in 5’ – 3’
direction
Leading strand synthesis
Template DNA: 3’ – 5’
orientation
New DNA: continuously
synthesized in 5’-
3’ direction
towards the
replication fork
Lagging strand synthesis
Template DNA: 5’ – 3’
orientation
New DNA: discontinuously
synthesized in 3’-
5’ orientation
away from the
replication fork
Okazaki fragments – are short, discontinuous replication products that are
produced in the lagging strand
- named after scientists Reiji Okazaki and Tsuneko
Okazaki
Step 6: DNA polymerase I uses its 5’ – 3’ exonuclease activity to
remove the RNA primer and fill the gap with new DNA.

Step 7: DNA ligase joins the ends of DNA fragments together

TERMINATION
Step 8: The two replication forks meet
Step 9: Once replication is complete,
the two double-stranded circular DNA
molecules (daughter strands) remain
interlinked. Topoisomerase II makes
double stranded cuts to unlink these
molecules.
DNA Replication in Eukaryotes
CELL CYCLE
Stage Activity Duration

Growth and
G1 increase in 10 hr
cell size

S DNA synthesis 8 hr
Post-DNA
G2 5 hr
synthesis
M Mitosis 1 hr
The steps for DNA replication are generally the same for all prokaryotic
and eukaryotic organisms

- New DNA strands begin with small primers of RNA

- Helicase unwinds the DNA
- DNA polymerase manufactures new DNA strand
- Semi conservative replication (leading and lagging strand
produced)
- Okazaki fragments produced
- DNA ligase seals any gaps
Differences between Prokaryotic and Eukaryotic
DNA Replication
EUKARYOTIC CELL PROKARYOTIC CELL
25x more DNA than prokaryotes Less DNA
Has multiple points of origin Has only one point of origin
Replication happens in the nucleus Replication happens in the
cytoplasm
Replication happens at a slower Replication happens at a faster
rate than prokaryotes rate than eukaryotes

Replication occurs during the S Replication occurs almost

phase only of the cell cycle continuously