DNA repair is crucial to maintaining low mutation rates between generations. There are several pathways for repairing damaged DNA: Mismatch repair detects and fixes errors made during DNA replication that escaped proofreading. In E. coli, the MutS and MutL proteins recognize mismatches and recruit enzymes to excise and resynthesize the damaged strand. Photoreactivation directly reverses UV radiation damage by breaking bonds between pyrimidine dimers using light energy. Base excision repair removes damaged bases using glycosylases that flip the base out of the double helix to cut it from the backbone. Polymerases and ligases then restore the intact strand. Nucleotide excision repair recognizes distortions in the

1. MOLECULAR BIOLOGY
Topic: DNA REPAIR
PRESENTED BY,
TARANJEET KAUR
B.Sc. BIOTECHNOLOGY
ROLL NO: 06
UNDER THE GUIDANCE OF,
Dr. SANTOSH KUMAR SINGH
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2. DNA Repair.
• DNA repair is a collection of processes by which a cell identifies
and correct the damage to the DNA molecule.
• The perpetuation of the genetic material from generation to
generation depends on maintaining rates of mutation at low
levels.
• High rates of mutation in the germ line would destroy the
species.
• Two important sources of mutation are inaccuracy in DNA
replication and chemical damage to the genetic material.
• Replication errors arise due to tautomerization, which imposes
an upper limit on the accuracy of base-pairing during DNA
replication.
• The enzymatic machinery for replicating DNA attempts to cope
with the mis-incorporation of incorrect nucleotides through a
proofreading mechanism, but some errors escape detection.
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3. • The natural and unnatural chemicals and radiation, break the
backbone of DNA molecule and alter its bases.
• Thus, errors in replication is unavoidable.
• Third important source of damage is the insertion of
Transposons.
• Errors in replication and damage to DNA have two
consequences. One is, permanent changes to the DNA, which
alters the coding sequences of a gene. Second is, the damage to
the DNA molecule prevent it’s use as a coding template for
replication and transcription.
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4. Mismatch Repair Removes Errors That
Escape Proofreading.
• Final responsibility for the fidelity of DNA replication lies with
mismatch repair system, which increases the accuracy of DNA
synthesis by an additional two or three orders of magnitude.
• Mismatch repair system faces two challenges. First, it must scan
the genome for mismatches, because mismatch are transient
and the mismatch must rapidly find and repair mismatches.
Second, the system must correct the mismatch accurately; that
is, it must replace the misincorporated nucleotide in the newly
synthesized strand and not the correct nucleotide in the parental
strand.
• In E.coli mismatches are detected by a dimer of the mismatch
protein MutS. MutS scans the DNA, recognizing mismatches
from the distortion caused in DNA backbone.
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5. • MutS embraces the mismatch containing DNA, inducing a
pronounced kink in the DNA, and a conformational changes in
itself.
• This complex of MutS and the distorted DNA recruits MutL, a
second protein component.
• MutL in turns activates MutH, an enzyme that causes an incision
or nick on one strand near the site of the mismatch.
• Nicking is followed by the specific helicase(UrvD) and one of the
three exonuleases.
• Helicase unwinds the DNA, starting from the incision and
moving in the direction of the site of the mismatch, and the
exonuclease progressively digests the displaced single strand.
This produces a single stranded gap, which is then filled by DNA
polymerase III and sealed with DNA ligase.
• The E.coli enzyme Dam methylase methylates A residues on
both the strands of the sequence 5’-GATC-3’. This sequence is
widely distributed along the entire genome.
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6. • When a replicatiion fork passese through the DNA that is
methylated at GATC on both strands, the resulting daughter DNA
duplex will be hemimethylated. Thus, for a few minutes, until
the Dam methylase catches up and methylates the newly
synthesised strand, daughter DNA duplexes will be methylated
only on the strand that served as a template. The newly strand is
thus marked and recognized as the strand for repair.
• Different exonucleases are used to remove single-stranded DNA
between the nick created by MutH and the mismatch,
depending on whether MutH cuts the DNA on the 5’ or the 3’
side of the misincorporated nucleotide. If the DNA is cleaved on
the 5’ side of the mismatch, the exonuclease VII or RecJ, which
degrade DNA in a 5’ to 3’ direction, remove the stretch of DNA.
Conversely if the nick is on the 3’ side of the mismatch, then
DNA is removed by exonuclease I, which degrades DNA from 3’
to 5’ direction.
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7. ARKA JAIN UNIVERSITY 7

8. Photoreactivation: Direct reversal of DNA
damage.
• Photoreactivation directly reverses
the formation of pyrimidine dimers
that results from ultraviolet radiations.
• In photoreactivation, the enzyme DNA
photolyase captures energy from light
and uses it to break the covalent
bonds linking adjacent pyrimidines.
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9. Base Excision Repair Enzymes Remove
Damaged Bases by a Base-Flipping Mechanism.
• In the base excision repair, an enzyme called a glycosylase
recognizes and removes the damaged base by hydrolizing the
glycosidic bond.
• The resulting abasic sugar is removed from the DNA backbone in
a further endonucleolytic step.
• Dna glycosylase flip the damaged base out of the double helix
and cleave theN-glycosidic bond of the damaged base, leaving
the abasic site.
• After the damaged nucleotide has been entirely removed from
the backbone, a repair DNA polymerase and DNA ligase restore
an inact strand using the undamaged strand as a template.
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10. • DNA glycosylase diffuse laterally along the the minor groove of
the DNA until a specific kind of lesion is detected. X-ray
crystallographic studies reveal that the damaged base is flipped
base is flipped out so that it projects away from the double
helix, where it sits in the specificity pocket of the glycosylase.
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11. Nucleotide Excision Repair Enzymes Cleave
Damaged DNA on either side of the lesion.
• Nucleotide excision repair enzymes do not recognize any
particular lesions, rather this system works by recognizing
distortions to the shape of the double helix, such as those
caused by thymine dimer or by the presence of a bulky chemical
adduct on a base. Such distortions trigger a chain of events that
lead to the removal of a short single-stranded segment that
includes the lesion.
• This removal creates a single-stranded gap in the DNA, which is
filled in by DNA polymerase using the undamaged starnd as a
template.
• Nucleotide excision repair in E.coli is largely accomplished by
four proteins: UrvA, UrvB, UrvC, and UrvD.
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12. • A complex of UrvA and UrvB scans the DNA, with UrvA being
responsible for detecting the distortions of the helix.
• Upon encountering distortions, UrvA exists the complex and
UrvB melting the DNA to create a single stranded bubble around
the lesion.
• Next, UrvB recruits UrvC, and UrvC creates two incisions: one
located eight nucleotides away on the 5’ side of the lesion and
the other four or five nucletides away on the 3’ side of the
lesion.
• These cleavages create a 12 to 13 residue long single stranded
DNA segment, which is made accessible by the action of the
DNA helicases UrvD.
• Finally, DNA polymerase I and DNA ligase fill in the resulting gap.
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13. ARKA JAIN UNIVERSITY 13

14. THANKYOU
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