UNIVERSITY OF SANTO TOMAS

FACULTY OF PHARMACY
DEPARTMENT OF BIOCHEMISTRY

DNA REPLICATION
PREPARED BY:
AARON | ADDUN | AGUILAR | AQUINO | BADUA |
BELARDO | CABASAG | CARLOS | CONDE
3A - BIOCHEMISTRY
INTRODUCTION: REPLICATION
• Part of Central Dogma of Molecular Biology
DNA RNA Proteins
• Copying of DNA
• A process in which genetic information is preserved and transmitted
to new cells and offspring
• Semi-conservative
• Orderly and sequential
• Highly accurate
• The process is catalyzed by specific enzymes
• Three Phases: initiation, elongation, and termination
DEFINITION OF
` TERMS
o Lagging strand
o DNA Replication
o Okazaki Fragments
o Semiconservative Replication
o DNA ligase
o Conservative Replication
o Helicase
o Dispersive Replication
o DNA Polymerase III
o Replication forks
o Exonuclease
o Temperature-Sensitive mutants
o Spontaneous mutation rate
o DNA Gyrase (Topoisomerase II)
o Replicon
o DNA Polymerase
o ARS ( Autonomous Replicating
o RNA Primer Sequences)
o Semidiscontinuos Replication o ORC (Origin Recognition Complex
o Leading strand o Replication foci
DNA
Replication
3 possible
models
DNA REPLICATION
INITIATION
• Replications starts at the ORIGIN
- initiator proteins identify specific base
sequences on the DNA called the ORIGIN SITE

• Prokaryotes – single origin site

• Eukaryotes – multiple sites of origin

• Polymerization ALWAYS occurs in a 5’-3’ direction

INITIATION
 INITIATION
• UNWINDING PROCESS
• DNA Helicase unwinds the dsDNA
• Single-stranded DNA-binding proteins (SSBs) stabilize the
complex
• Topoisomerase II (DNA Gyrase) relieve torsional stress by ATP
Hydrolysis
 ELONGATION
• Semi-discontinuous Replication
Anti parallel strands are replicated simultaneously
 Leading strand is synthesized continuously
 Lagging strand is synthesized into fragments (discontinuously)
 ELONGATION
• RNA Primers are required
 ELONGATION
• The two DNA polymerases can replicate both strands by
causing the newly synthesized lagging strand to loop out
between the polymerase and the fork. Once the
polymerase completes an Okazaki Fragment, it
dissociates from the DNA template.
• A new primer is then produced at the replication fork, the
polymerase re-associates with the template to continue
synthesis of the lagging strand.
A. C.

B. D.

E.
ELONGATION
 TERMINATION

• 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.
CORE PROTEINS AT THE REPLICATION FORK

Topoisomerases - prevents torsion by DNA breaks

Helicases - separates 2 strands
Primase - RNA primer synthesis
Single strand - prevent reannealing of single
binding proteins strands
DNA polymerase - synthesis of new strand
DNA ligase - seals nick via phosphodiester
linkage
SUMMARY
• Initiation
• Proteins bind to DNA and open up double helix
• Prepare DNA for complementary base pairing
• Elongation
• Proteins connect the correct sequences of
nucleotides into a continuous new strand of DNA
• Termination
• Proteins release the replication complex
EUKARYOTIC REPLICATION
REVIEW OF RELATED LITERATURE
Targeting DNA Replication Stress for
Cancer Therapy
1, 1 2
By Jun Zhang *, Qun Dai , Dongkyoo Park and Xingming
2,
Deng *

2016

Reference: Zhang, J. et.al. (2016). Targeting DNA replication

stress for cancer therapy. Retrieved January 24, 2017 from:
http://www.mdpi.com/2073-4425/7/8/51c
SUMMARY OF RRL

• The aim of this study was to:

• Understand the DNA replication process and how
the DDR (DNA damage response) of normal cells
differed from cancer cells.
• Identify approaches that enhance replication stress,
which leads to death of cancer cells.
 KEYPOINTS
• Low levels of replication stress promotes genome
instability and fuels tumorigenesis. High levels of
replication stress can cause death of cancer cells.

• Radiation - Direct DNA damage and interferes with

DNA replication.

• Chemotherapeutic agents - Induce stalling of

replication forks.
KEYPOINTS
• Inhibitors - Induces fork stalling and collapse, which leads to cell death.

• Ex: ChK1 inhibitors

“DNA replication is inappropriately initiated from multiple origins, leading to the

exhaustion of replication factors and to fork stalling and collapse.” (Zhang, J. et.al.

,2016)

“Excessive replication stress is deleterious even in cancer cells as they cannot

complete mitosis with unreplicated regions of genome, and will undergo mitotic

catastrophe if replication stress persists into mitosis” (Zhang, J. et.al. ,2016)