DNA Replication and Repair

Prof. DR. dr. Hadyanto Lim, M.Kes, SpFK, FESC, FIBA,

FAHA
Department of Pharmacology and Molecular Sciences
Faculty of Medicine, Methodist University of Indonesia - Medan
Molecular Biology Research, Postgraduate School,
University of Sumatra Utara - Medan

Why DNA replication is

needed ?
DNA inside the cells is
damaged by
Chemicals
Radiation
Thermal
Reactive molecules
generated inside the cell

Levels of Reproduction
Organism : Sexual and
asexual
Cells : Cellular division
Genetic
: DNA replication

DNA
Replication

Meselson Stahl Experiment

Study Design
DNA extracted
from bacteria
mixed with a
concentrated of
the salt cesium
chloride (CsCl),
then centrifuged
until the doubled
DNA molecules
reach equilibrium
according to their
density.
The density
formed with the
lowest at the top
and highest at the
bottom.

Results of the density-gradient experiments obtained by

Meselson Stahl demonstrates that replication occurs
semiconservatively (by Watson and Crick)

The appearance of
the hybrid band and
the disappearance of
the heavy band after
one generation
eliminates
conservative
replication
The subsequent
appearance of two
bands, one light and

DNA double helix acts as a template for its

own duplication

The chemistry of
DNA synthesis

DNA synthesis catalyzed by DNA

polymerase

Basic Requirements in DNA

Synthesis
A single-stranded DNA circle cannot serve as a
template for DNA polymerase because the
enzyme
cannot initiate the formation of a DNA strand.
Rather, it can only add nucleotides to the 3
hydroxyl terminus of an existing strand. The
strand that provides the necessary 3OH
terminus
is called primer.
Both prokaryotic and eukaryotic have the same
two basic requirements : a template DNA strand
to copy and a primer strand to which nucleotides
can be added.

Question
How the DNA replication
proceeds in prokaryotes
and eukaryotes ?

Replication in Bacteria Cells

Two replication forks moving in opposite directions on a

circular chromosome

DNA Repair
Correcting the spontaneous
changes in DNA.
Changes could be due to heat,
metabolic accidents, radiation,
exposure to substances in the
environment.
Fewer than one in 1000 accidental
base changes in DNA results in a
permanent mutation.

Xeroderma Pigmentum

DNA Repair
Most repair system require that a
damaged section of the DNA be
excised, that is, selectively
removed.
Consequently, if one or more
nucleotides is removed from one
strand, the complementary strand
can serve as a template for
reconstruction of the duplex.

Nucleotide Excision
Repair (NER)
Operates by a cut-and patch
mechanism that removes a variety
of bulky lesions, including
pyrimidines dimers and
nucleotides which various chemical
groups have become attached.

A Pyrimidine dimer that has formed within a DNA duplex

following UV irradiation

Failure to repair such

lesions produces
permanent alterations, or
mutations, in the DNA
-Gametes
-Somatic Cells

Nucleotide Excision
Repair (NER)
Two pathways distinguished in NER
- Transcription-coupled
pathway , repairs the
template)
- Global pathway, corrects DNA
strands in the
remainder of the genome)

Nucleotide excision
repair
(Global
Pathway)
Separation
(Pemisahan)
Incision
(Pemotongan)
Excision
(Pembuangan)
Filled by DNA
polymerase

Sealed (Penempelan)
by DNA
ligase

Nucleotide Excision Repair (NER) (Global

Pathway)

Damage recognition in the globa

is mediated by an XPC protein
damage recognition in the tran
coupled pathways is thought to
mediated by a stalled RNA poly
in conjunction with a CSB prot

DNA strand separation (by XPB a

proteins, two helicase subunits

Nucleotide Excision Repair (NER)

(Global Pathway)

Incision (by XPG on the

3side and the XPF-ERCC1
complex on the 5side) by a
pair of endonucleases. (3)

Excision (4)

DNA repair synthesis (by

DNA Polymerase and/or ).
(5)

Ligation by DNA ligase I (6)

The End