Transcript The Mutability and Repair of DNA

The Mutability and Repair of
DNA
Replication errors and their repair
The nature of mutations: Point mutation
1. Switch of one base for another:
(transition)
(transversion)
purine
pyrimidine
2. insertion or deletion of a nucleotide
Drastic changes in DNA
Deletion
Insertion
Rearrangement of chromosome
By insertion of a transposon, or aberrant actions of recombination
Process.
Some replication errors escape proofreading
Mismatch repair removes errors escape proofreading
1. It must scan the genome.
2. The system must correct the mismatch accurately.
Scan DNA
Distortion in the backbone
MutL activate MutH
Embracing mismatch;
Inducing a kick in DNA;
Conformational change in
MutS itself
Nicking is followed by Helicase (UvrD) and one of exonucleases
(III)
DNA methylation to recognize the parental strain
Once activated,
MutH selectively nicks the
Unmethylated strand.
Directionality in mismatch repair
Mismatch repair system in Eukaryotics
E. coli
Eukaryotics
MutS
MutL
MSH
MLH or PMS
(MutS homolog)
Hereditary nonpolyposis colorectal cancer
(mutations in human homologes of Muts and MutL)
DNA damage
Radiation, chemical mutagens, and spontaneous damage
spontaneous damage due to hydrolysis and deamination
Deamination converts adenine to
hypoxanthine, base pair with C
Deamination converts Guanine to
xanthine, base with C but only two H
bonds
deamination
Base pair with A
depurination
DNA damage
spontaneous damage to generate natural base
deamination
Methylated Cs are hot spot for spontaneous mutation in vertebrate DNA
Damaged by alkylation and oxidation
Alkylation at the oxygen of carbon atom 6 of G : O6-metylguanine,
often mispairs with T.
Oxidation of G generates oxoG, it can mispair with A and C. a
G:C to T:A transversion is one of the most common mutation in
human cancers.
Gamma radiation and X-rays
• Cause double-strand breaks in the DNA,
which are difficult to repair.
• Ionizing radiation and agents like
bleomycin that cause DNA to break are
said to be clastogenic (p245).
DNA damage by UV
Thymine dimer
These linked bases are incapable of base-pairing and cause
DNA polymerase to stop.
Mutations caused by base analogs and intercalating agents
Base analogs
Thymine analog
Analogs mispair to cause mistakes during replication
Mutations caused by intercalating agents
Intercalating agents
flat molecules
Causing addition or deletion of bases during replication
Repair of DNA Damage: DNA repair system
Excision repair systems: the damaged nucleotide is not repaired but removed from
the DNA, the other undamaged strand serves as a template for reincorporation of
the correct nt by DNA polymerase
Recombination repair: both strands are damaged. Sequence information is
retrieved from a second undamaged copy of the chromosome.
Direct reversal of DNA damage
photoreactivation
Capture energy from light
breaking
covalent bond
O6-metylguanine
To its own cytosine
Base excision repair
DNA glycosylases are lesion-specific and cells have multiple DNA glycosylases
1. Uracil glycosylase
2. Another specific glycosylase is responsible for removing oxoG
AP: apurinic or apyrimidinic
Base excision repair
If a damaged base is not removed by base excision before DNA
replication: a fail-safe system
oxoG:A repair
Nucleotide Excision Repair
Recognizing distortions to the shape of the DNA
(thymine dimer or bulky chemical adduct)
UvrA detecting distortion
8 nt away from 5’
4-5 nt away from 3’
UvrB melting DNA
In E.coli: 4 proteins involved
Nucleotide Excision Repair
The principles of nucleotide excision repair in higher cells is much the
same as in E. coli but us moer complicated, involving 25 or more
polypeptides.
The UVR proteins are needed to mend damage from UV light;
Mutants of uvr genes are sensitive to UV light, and lack the capacity to
remove T-T or T-C adducts.
In human, xeroderma pigmentosum patients have mutations in seven genes
(XP genes). These XP proteins are corresponding to proteins involved in
nucleotide excision repair.
Transcription-coupled repair
Involves recruitment to the stalled
RNA polymerase of nucleotide
excision repair proteins
It focuses repair on genes being
actively transcribed.
TFIIH unwinds the DNA template
during the initiation of
transcription. Subunits of TFIIH
include the DNA helix-opening
proteins XPA and XPD.
Translesion DNA synthesis: although are template dependent, the
synthesis in a manner that is independent of base pairing
obstacles to progression of the DNA polymerase
(or AP site)
Complex of proteins UmuC and D’
(Y-family of DNA polymerase)