1. The Mutability and Repair of DNA

2. Replication errors and their repair
The nature of mutations: Point mutation
1. Switch of one base for another:
purine
pyrimidine
(transition)
(transversion)
2. insertion or deletion of a nucleotide

3. Rearrangement of chromosome
Drastic changes in DNA
Deletion
Insertion
Rearrangement of chromosome
By insertion of a transposon, or aberrant actions of recombination
Process.

4. Some replication errors escape proofreading

5. 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)

6. DNA methylation to recognize the parental strain
Once activated,
MutH selectively nicks the
Unmethylated strand.

7. Directionality in mismatch repair

8. Mismatch repair system in Eukaryotics
MutS
MutL
E. coli
MSH
(MutS homolog)
MLH or PMS
Eukaryotics
Hereditary nonpolyposis colorectal cancer
(mutations in human homologes of Muts and MutL)

9. 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

10. DNA damage spontaneous damage to generate natural base
deamination
Methylated Cs are hot spot for spontaneous mutation in vertebrate DNA

11. 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.

12. 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).

13. DNA damage by UV Thymine dimer
These linked bases are incapable of base-pairing and cause
DNA polymerase to stop.

14. Mutations caused by base analogs and intercalating agents
Thymine analog
Analogs mispair to cause mistakes during replication

15. Intercalating agents Mutations caused by intercalating agents
flat molecules
Causing addition or deletion of bases during replication

16. 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.

17. Direct reversal of DNA damage
photoreactivation
Capture energy from light
breaking
covalent bond
To its own cytosine
O6-metylguanine

18. Base excision repair AP: apurinic or apyrimidinic
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

19. Base excision repair
If a damaged base is not removed by base excision before DNA replication: a fail-safe system
oxoG:A repair

20. Nucleotide Excision Repair
Recognizing distortions to the shape of the DNA
(thymine dimer or bulky chemical adduct)
4-5 nt away from 3’
UvrA detecting distortion
8 nt away from 5’
UvrB melting DNA
In E.coli: 4 proteins involved

21. 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.

22. 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.

23. 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)