Presented by

Nashmiya Saheer
 DNA damage if left unrepaired can lead
to mutations
 DNA damage is not the same as the
mutation
 DNA damage is a chemical alteration to
DNA
 While mutation is a change in base pair
 DNA can be damaged by different
environmental factors,mutagens and it
can alse happen spontaneously.
1) Hydrolysis and deamination
2) Gamma and X-rays
3) Ultraviolet radiation
4) Alkylation of bases
5) Oxidation
6) Base analogues
7) Intercalating agents
 Most important hydrolytic damage is deamination

C =G A=T

A=T C=G

G=C C-G

C=G T=A
 Mutational Hotspots
vertebrate DNA contain 5-
methylcytosine in place of
cytosine.Deamination of 5-methyl cytosine
yeild thymine which is a normal base of
DNA which will not be recognized as an
abnormal base.Thus methylated cytosines
are known as hotspots for spontaneuos
mutations.
 X-rays and gamma rays like uv rays can directly interact with DNA
 They cause damage by ionizing molecules especially water
surrounding DNA
 This forms chemical substances with an unpaired electron known
as free radicals
 These free radiclas especially those containing oxygen are very
reactive and attacks DNA,change a base,but frequently causes a
single or double strand break
 Ss breaks are not serious and easily repaired.
 But ds breaks are difficult to repair and causes a lasting mutation
 Because ionizing radiation can break chromosome it is known as
clastogen which means “breaker”
 Uv rays cross links adjacent pyramidines in same strand forming
pyramidine dimers
 These dimers block DNA replication because the machinary
cannot decide which bases to insert opposite the dimer
 Radiation of wavelength 260nm is strongly absorbed by bases
 In the case of thymine dimers,they consist of a cyclobutane ring
formed by links between carbon atoms of adjacent thymines
 The process of addition of alkyl groups (carbon
containing group)to the negative centres of DNA
by electrophilic substances is known as
alkylation
 The favorite site of attack is N7 of guanine and N3
of adenine
 N7 alkylation of guanine does not change the
base and is harmless
 N3 alkylation of adenine yield 3-methyl adenine
which cannot base pair with any other
base,hence noncoding base.
 Thus DNA polymerase stops replication when it
encounters a 3-methyl adenine.
 • The alkylation target O6 guanine leads to most
mutations
• it allow the product to base pair with thymine rather
than cytosine
• A laboratory mutagen ethylmethane
sulfonate(EMS)transfer ethyl group to DNA
• Alkylation of O6 guanine changes the tautomeric form
of guanine so it base pair with thymine
• This leads to GC to AT base pair transition
 oxidation
 Reactive oxygen species O2-,h2O2,OH are generated by
ionizing radiations and chemical agents.
 Oxidation of guanine yield 7,8-dihydro-8-oxoguanine or
OXOG
 OXOG adduct is highly mutagenic because it can base pair
with both adenine and cytosine resulting in G:C To T:A
transversion
Base analogs
 Structurally similar to normal bases
 They will be taken up by the cell converted into nucleoside
and incorporated into DNA during replication
 But due to the structural differences with normal base they
base pair inaccurately leading to DNA damage
 Eg ; 5-bromouracil an analogue of thymine
 Molecules containing polycyclic rings
 Bind to purins or pyramidines just as the
bases bind with each other
 Eg :proflavin,acridine,ethidium
 Causes deletion or addition of bases
 Leading to shift in coding sequence
2 consequences of DNA damage are :
1. Creates error in replication or
transcription
2. Create altered bases that cause
mispairing
 Thus cells have to evolve repair
mechanisms to identify and repair
damage before mutation
 Mismatch repair sytem monitor structural defects
rather than chemical changes and is more sensitive
 This system cuts out part of the DNA strand
containing the wrong base
 The gap is then filled by DNA polymerase III
 To repair the wrong base,the mismatch system need
to know which is the new strand
 In E.coli the chromosome is mathylated by 2 system:
1. Dam(DNA adenline methylase) – converts adenine
in GATC to 6-methyladenine
2. Dcm(DNA cytosine methylase) – converts cytosine
in CCAGG and CCTGG to 5-methyl cytosine
 The Dam and Dcm methylase take few
minutes to methylate the new strand
 During this period DNA will be hemi
methylated
 During this period variety of repair
systems check the DNA for mismatches.
 Another important mismatch repair
system is mutSHL repair system in E.coli
 Also known as cut and patch repair
system
 Recognizes bulges in DNA
 Mostly repair UV damages such as
thymine dimers
 Defects in this repair system results in
lowered resistance to UV
 Hence this system is known as Uvr
 These specific repair mechanisms identify specific chemical
change sin DNA
 Deamination results in :
A. hypoxanthine - adenine
B. xanthine - guanine
C. uracil - cytosine
 DNA glycosylases removes the unnatural bases by breaking
the bond between the base and deoxyribose sugar
 Specific glycosyases exist for different bases.for eg-uracil-
N-glycosylase or ung protein remives uracil
 Removal of bases resuts in apurinic or apyrimidic site (AP
site)
 Then endonuclease cuts next to AP site
 A free 3’OH is created and DNA polymerase I makes new
DNA
 The final nick sealed by DNA ligase
 Allowing DNA replication to occur through severely
damaged area is known as SOS error prone repair
 Single stranded damaged DNA activated Rec A
protein that induces the SOS system
 SOS system is repressed by Lex A which will be self
cleaved by the induction caused by Rec A protein
 Important SOS genes are :
1. umuC (umu=ultraviolet mutagenesis)
2. umuD
 These 2 code for polymerase V which can replicate
past pyrimidine dimers and missing bases
 Also known as post replication repair
 Replication past pyrimidine dimer creates a problem
since the replication machinary stops at such dimers
 After a [pause replicatiom continues leaving a gap in
the new strand
 Next recombination occurs between the gapped
strand and its homologue in the other daughter DNA
duplex
 This recombination depend on Rec A protein
 The net effect of this recombination is to fill the gaps
in the strand and to create a new gap in the other
DNA
 Because the other DNA has no dimer the gap can be
easily filled by DNA polymerase and ligase
 The enzyme DNA photolyase capture
energy from light and uses it to
break the covalent bond linking the
thymine dimers
 It occurs only when visible light is
present
 Ifthe template strand of a DNA is damaged
that is being transcribed,RNA polymerase
may grind to a halt.such damage is repaired
by transcription coupled exicision repair
 Transcription repair coupling factor,TRCF
detects stalled RNAP and directs UvrAB
proteins to the site of block,
 TRCF detaches the polymerase so it does
not hinder UvrAB proteins
 Deamiation of 5-methyl cytosine yield
thymie which is a natural base
 Causes a T-G mismatch
 A specific endonuclease nicks the DN next
to the T of T-G mismatch
 Also known as very short patch repair
 Nicking enzyme is known as Vsr
endonuclease
 Then DNAP I removes a short length of DNA
and replaces with a new strand of DNA
Some methylated groups
attached to oxygen require
sepcial treatment
They are removes by suicidal
Protiens
These proteins attaches
Methyl group to themselves
And get degraded