DNA Repair Flashcards
What are the 2 main types of DNA damage?
Chemical alteration and spontaneous damage.
What is meant by exogenous chemical alteration?
Damage caused by environmental mutagens such as UV radiation.
What is meant by endogenous chemical alteration?
Damage caused by internally generated mutagens such as hydroxyl radicals which as formed as by products of metabolic reactions.
Give examples of spontaneous damage to DNA.
Deamination and depurination.
How does UV radiation damage DNA?
Induces formation of pyrimidine dimers where two adjacent pyrimidines are joined by a cyclone take ring. Causes loss of aligned C=C bonds, loss of planarity and distortion of the DNA structure.
How does alkylation damage DNA?
Addition of methyl/ethyl groups to various positions on the DNA bases.
How do carcinogens damage DNA?
React with DNA bases resulting in the addition of large bulky chemical groups to the DNA molecule. Many carcinogens are activated via reactions with cytochrome P450, this means that non-toxic drugs may become toxic once processed.
How often does depurination of A/C/G occur?
Approximately 500 times a day.
What occurs as a result of depurination of A/C/G?
Changes base pair properties and causes mismatches.
How often does depurination of A/G occur?
Approximately 18,000 times a day.
What occurs as a result of depurination of A/G?
Cleavage of the bond between the purine base and deoxyribose- elimination of the base.
Which is more damaging deamination or depurination?
Depurination- is more toxic and more mutagenic.
What are the two main methods of DNA repair? And which is used most often.
Direct reversal- rare, excision repair- more likely.
How are pyrimidines dimers repaired?
Visible light is used by a photo reacting enzyme to break the cyclobutane.
Where is pyrimidine dimer direct reversal seen?
In e.coli, yeast and some plants.
How is guanine methylation repaired?
O6-methylguanine methyltransferase has a Cys residue that is methylated by the methylguanine. This is a suicide enzyme.
What occurs during excision repair?
Damaged bases are removed and replaced with newly synthesised DNA.
Describe base-excision repair.
DNA glycosylase cleaves glycosidic bond between base and deoxyribose. Leaves an AP site and the backbone intact. AP endonuclease cleaves DNA chain. Deoxyribosephosphodiesterase removes remaining deoxyribose. Gap filled by DNA Pol and sealed by ligase.
Describe nucleotide excision repair.
3’ and 5’ endonucleases cleave the backbone upstream and downstream of a bulge cause by a thymine dimer. Helicase unwinds DNA, causing excision of a 30nt oligonucleotide containing the thymine dimer. Gap is filled by DNA Pol and sealed by ligase.
What enzymes catalyse nucleotide excision repair in E.coli and what do mutations in these cause?
The uvrABC gene productions. Mutations lead to high UV sensitivity.
What enzymes catalyse nucleotide excision repair in eukaryotes and what do mutations cause?
The RAD gene products. Mutations cause Xeroderma pigmentosa.
Describe Xeroderma pigmentosa.
Individuals cannot repair DNA damage caused by UV exposure. RARE genetic disease. Results in reduced life expectancy due to high levels of skin cancer if exposed to UV light. Sufferers must be completely shielded from UV radiation.
When is the mismatch repair mechanism used?
To repair post-replication mismatches.
Which is the only repair mechanism to increase fidelity during DNA replication?
Mismatch repair
What proteins are involved in mismatch repair in E.coli?
MutHLS
What is the role of MutS?
Recognises mismatch
What is the role of MutL?
Binds MutS at mismatches and recognises hemimethylated dam sites. Uses ATPase activity to translocate along the DNA looking for a hemimethylated dam site.
Why is DNA repair a crucial process?
Information must be passed on from one generation to the next and must be stable over many lifetimes. DNA must be repaired to limit information loss.
What is the role of MutH?
- get from flashcard
Describe the process of mismatch repair in E.coli.
MutS recognises mismatch and MutL binds. MutH is activated when bound to MutL and cleaves the non-methylated strand opposite the hemimethylated dam site. This defines the newly synthesised strand for repair. UvrD helicase excises everything between the mismatch site and the excision site. Gap filled by DNA Pol III and sealed by ligase.
Where is the nick produced by MutH?
Can be upstream or downstream of the mismatch- affects which endonuclease is needed for excision.
What catalyses mismatch repair in mammalian cells?
MSH complex.
How are daughter strands distinguished from parental strands in mammalian cells?
Excision occurs between Okazaki fragments and the mismatch site.
Which part of the replication machinery remains bound to the DNA during mismatch repair in mammalian cells?
beta clamps- remain bound and interact with repair proteins.
What can be the result of mutations in the MSH complex?
Non-polyposis colorectal cancer.
