DNA Repair Mechanisms

Living organisms contain many enzymes that scan

their DNA for damage and initiate repair processes
when damage is detected.

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 DNA Repair Mechanisms in E. coli

1. Light-dependent repair (photo-reactivation)

2. Excision repair
3. Mismatch repair
4. Post-replication repair
5. Error-prone repair system (SOS response)

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1) UV Light-Dependent
Repair: Photolyase
Cleaves Thymine
Dimers.

--No Endonuclease
--No Ligase

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 2) Types of Excision Repair
• Base excision repair pathways remove
abnormal or chemically modified bases.

• Nucleotide excision repair pathways remove

larger defects, such as thymine dimers.

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 Excision Repair
• A DNA repair endonuclease or endonuclease-
containing complex recognizes, binds to, and
excised the damaged base or bases.

• A DNA Polymerase fills in the gap, using the

undamaged complementary strand of DNA as a
template.

• DNA ligase seals the break left by DNA

polymerase.

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 Base Excision Repair

AP:apyrimidinic site
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( )

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Nucleotide Excision Repair

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 3) Mismatch Repair in E. coli
• The mismatched nucleotide is excised from the new
strand and replaced with the correct nucleotide, using the
methylated parental strand as a template.

• Mismatching or mispairing of G and T

• The A in GATC sequences is methylated subsequent to

DNA replication.

• In newly replicated DNA, the parental strand is

methylated, but the new strand is not. This difference
allows the mismatch repair system to distinguish the new
strand from the old strand.
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• MutS recognizes
mismatches and binds to
them to initiate the repair
process.
• MutH and MutL join the
complex.
• MutH cleaves the
unmethylated strand at
hemimethylated GATC
sequences on either side of
the mismatch.
• Excision requires MutS,
MutL, MutU (DNA helicase
II), and an exonuclease.

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• DNA polymerase III
fills in the gap, and
DNA ligase seals
the nick.

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 4) Post-replication Repair in E. coli
• A thymine dimer in the template strand blocks replication
(DNA Polymerase III does not recognize thymidine dimer)

• DNA Polymerase III restarts DNA synthesis past the dimer,

leaving a gap in the nascent strand.

• RecA binds to the single strand of DNA at the gap and

mediates base pairing with the homologous segment of
the sister double helix to fill the gap.

• DNA polymerase fills the gap in the sister double helix,

and DNA ligase seals the nick.

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 5) The SOS Response in E. coli

• If DNA is heavily damaged by mutagenic agents, the SOS

response, which involves many DNA recombination, DNA
repair, and DNA replication proteins, is activated.

• DNA dependent DNA Polymerase V replicates DNA in

damaged regions, but sequences in damaged regions cannot
be replicated accurately.

• This error-prone system eliminates gaps but increases the

frequency of replication errors (Pol II, IV and V are low-
fidelity polymerases)

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 Induction of the SOS Response
• In the absence of DNA damage, LexA binds to DNA
regions that regulate transcription of SOS response genes
and keeps their expression levels low.
• When extensive DNA damage occurs, RecA binds to
single-stranded regions of DNA in damaged regions.
• This activates RecA, which stimulates LexA to inactivate
itself. When LexA is inactivated, the SOS response genes
are expressed.

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 Human Diseases with Defects in
DNA Repair
Several human disorders result from defects in
DNA repair pathways.

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Xeroderma Pigmentosum (XP)

• Individuals with XP are sensitive

to sunlight (UV light).

• The cells of individuals with XP are

deficient in the repair of UV-
induced damage to DNA.

• Individuals with XP may develop

skin cancer or neurological
abnormalities.

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 Mutations in Human Globin Genes
• Adult hemoglobin (Hemoglobin A) contains two 
chains and two  chains.

• Hemoglobin in patients with sickle-cell anemia

(Hemoglobin S) differs from Hemoglobin A at only
one position.

• The sixth amino acid in the  chain is glutamic acid in

Hemoglobin A (HBBA) and is valine in Hemoglobin S
(HBBS). This substitution is caused by mutation of a
single base pair (T:A substitution).

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 Tay-Sachs Disease

• Tay-Sachs disease is an
autosomal recessive
disease.
• The mutation causing
Tay-Sachs disease is in
the gene encoding
hexosaminidase A.

not linked to sex chromosome

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 Transgenic Mouse: Generic term for an engineered mouse that has a
normal DNA sequence for a gene replaced by an engineered sequence or a
sequence from another organism.

 Knockout Mouse: A transgenic mouse in which the normal gene is missing

or engineered so that is not transcribed or translated. “Knocks out” that
gene.

 Knockin Mouse: A transgenic mouse in which the engineered “transgene”

is subtly manipulated to: (A) alter the function of the gene (e.g., replace one
amino acid with another in a site to determine if that site is essential for the
protein’s function); (B) change transcription rate to overproduce or
underproduce the gene product; or (C) create a fluorescent gene product to
map its distribution in tissue.

 Conditional Knockout (Knockin) Mouse: A transgenic mouse in which

the transgene is knocked out (or in) in specific tissues, at a specific
developmental stage, or in response to an exogenous substance (e.g., an
antibiotic). 21