Lect-06

DNA damage, repair

and recombination
Learning Outcome
(LO)
After students learn this
lecture, students are able to
explain:
• Mutagenesis (LO 6.1)
• DNA damage (LO 6.2)
• DNA repair (LO 6.3)
• Recombination (LO 6.4)
 MUTAGENESIS
Mutation
❑ Mutations are heritable permanent changes in the base sequence of DNA.
❑ Point mutations may be transitions (e.g. G-C→A-T) or transversions (e.g. G-C→
T-A).
❑ Deletions and insertions involve the loss or addition of bases and can cause
frameshifts in reading the genetic code.
❑ Silent mutations have no phenotypic effect, while missense and nonsense
mutations change the amino acid sequence of the encoded protein.

LO 6.1: Students are able to explain mutagenesis

Biology _ concepts and investigations-McGraw-Hill (2012)

LO 6.1: Students are able to explain mutagenesis

MUTAGENESIS
Replication fidelity
❑ The high accuracy of DNA replication (one error per 1010 bases incorporated)
depends on a combination of proper base pairing of template strand and incoming
nucleotide in the active site of the DNA polymerase, proofreading of the
incorporated base by 3’→5’ exonuclease and mismatch repair.

LO 6.1: Students are able to explain mutagenesis

MUTAGENESIS
Physical mutagens
❑ Ionizing (e.g. X- and γ-rays) and nonionizing (e.g. UV) radiation produce a variety
of DNA lesions.
❑ Pyrimidine dimers are the commonest product of UV irradiation.

LO 6.1: Students are able to explain mutagenesis

MUTAGENESIS
Chemical mutagens
❑ Base analogs can mispair during DNA replication to cause mutations.
❑ Nitrous acid deaminates cytosine and adenine.
❑ Alkylating and arylating agents generate a variety of adducts that can block
transcription and replication and cause mutations by direct or, more commonly,
indirect mutagenesis.
❑ Most chemical mutagens are carcinogenic.

LO 6.1: Students are able to explain mutagenesis

MUTAGENESIS
Direct mutagenesis
❑ If a base analog or modified base whose base pairing properties are different from
the parent base is not removed by a DNA repair mechanism before passage of a
replication fork, then an incorrect base will be incorporated.
❑ A second round of replication fixes the mutation permanently in the DNA.

LO 6.1: Students are able to explain mutagenesis

MUTAGENESIS
Indirect mutagenesis
❑ Most lesions in DNA are repaired by error-free direct reversal or excision repair
mechanisms before passage of a replication fork.
❑ If this is not possible, an error-prone form of translesion DNA synthesis may take
place involving specialized DNA polymerases and one or more incorrect bases
become incorporated opposite the lesion.

LO 6.1: Students are able to explain mutagenesis

DNA DAMAGE
DNA lesions
❑ The chemical reactivity of DNA with exogenous chemicals or radiation can give
rise to changes in its chemical or physical structure.
❑ These may block replication or transcription and so be lethal, or they may generate
mutations through direct or indirect mutagenesis.
❑ The chemical instability of DNA can generate spontaneous lesions such as
deamination and depurination.

LO 6.2: Students are able to explain DNA damage

DNA DAMAGE
Oxidative damage
❑ Reactive oxygen species such as superoxide and hydroxyl radicals produce a
variety of lesions including 8-oxoguanine and 5-formyluracil.
❑ Such damage occurs spontaneously but is increased by some exogenous agents
including λ-rays.

LO 6.2: Students are able to explain DNA damage

DNA DAMAGE
Alkylation
❑ Electrophilic alkylating agents such as methylmethane sulfonate and
ethylnitrosourea can modify nucleotides in a variety of positions.
❑ Most lesions are indirectly mutagenic, but O6-alkylguanine is directly
mutagenic.

LO 6.2: Students are able to explain DNA damage

DNA DAMAGE
Bulky adducts
❑ Bulky lesions such as
pyrimidine dimers and
arylating agent adducts
distort the double helix
and cause localized
denaturation.
❑ This disrupts the normal
functioning of the DNA.

LO 6.2: Students are able to explain DNA damage

 DNA REPAIR
Photoreactivation
❑ Cleavage of the cyclobutane ring of pyrimidine dimers by DNA photolyases
restores the original DNA structure.
❑ Photolyases have chromophores which absorb blue light to provide energy for the
reaction.

LO 6.3: Students are able to explain DNA repair

 DNA REPAIR
Alkyltransferase

❑ An inducible protein specifically removes an alkyl

group from the O6 position of guanine and transfers
it to itself, causing inactivation of the protein.

LO 6.3: Students are able to explain DNA repair

 DNA REPAIR
Excision repair
❑ In nucleotide excision repair, an endonuclease makes nicks
on either side of the lesion, which is then removed to leave a
gap.
❑ This gap is filled by a DNA polymerase, and DNA ligase
makes the final phosphodiester bond.
❑ In base excision repair, the lesion is removed by a specific
DNA glycosylase.
❑ The resulting AP site is cleaved and expanded to a gap by an
AP endonuclease plus exonuclease.
❑ Thereafter, the process is like nucleotide excision repair.

LO 6.3: Students are able to explain DNA repair

 DNA REPAIR
Excision repair

LO 6.3: Students are able to explain DNA repair

 DNA REPAIR
Mismatch repair
❑ Replication errors which escape proofreading have a
mismatch in the daughter strand.
❑ Hemimethylation of the DNA after replication allows the
daughter strand to be distinguished from the parental strand.
❑ The mismatched base is removed from the daughter strand by
an excision repair mechanism.

LO 6.3: Students are able to explain DNA repair

 DNA REPAIR
Hereditary repair defects
❑ Mutations in excision repair genes or a translesion DNA
polymerase cause different forms of xeroderma pigmentosum,
a sun-sensitive cancer-prone disorder.
❑ Excision repair is also defective in Cockayne syndrome.

LO 6.3: Students are able to explain DNA repair

 RECOMBINATION
Homologous recombination
❑ The exchange of homologous regions between two DNA
molecules occurs extensively in eukaryotes during meiosis.
❑ In prokaryotes, recA-dependent recombination involves a
four-stranded Holliday intermediate which can resolve in two
ways.
❑ The integrity of DNA containing unrepaired lesions can be
maintained during replication by homologous recombination.

LO 6.4: Students are able to explain recombination

 RECOMBINATION
Homologous recombination

LO 6.4: Students are able to explain recombination

 RECOMBINATION
Site-specific recombination
❑ The exchange of nonhomologous regions of DNA at specific
sites is independent of recA.
❑ Integration of bacteriophage into the E. coli genome involves
recombination at a 15 bp sequence present in both molecules
and specific protein integration factors.
❑ Site-specific recombination also accounts for the generation of
antibody diversity in animals.

LO 6.4: Students are able to explain recombination

 RECOMBINATION
Transposition
❑ Replicated copies of transposable DNA elements can insert themselves anywhere
in the genome.
❑ All transposons encode a transposase which catalyzes the insertion.
❑ Retrotransposons replicate through an RNA intermediate and are related to
retroviruses.

LO 6.4: Students are able to explain recombination

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