Chapter 10 - The Mutability of Repair of DNA Flashcards
(33 cards)
Explain the difference between transition and transversion mutations.
Transistions - pyridine-to-pyrimidine and purine-ro-purine substitutions (T to G and A to G).
Transversions - pyrimidine-to-purine and purine-to-pyrimidine substitutions.
What are DNA microsatellites?
Mutation-prone sequences with di-, tri-, or tetra nucleotide sequences. Example: CA. Due to slippage of DNA polymerase.
CAG repeats cause polymorphism –> multiple extension causes polyglutamine stretches –> ex. Huntington’s disease.
Which protein detects mismatches in E. coli? How is this proteins specificity determined?
MutS. DNA containing a mismatch is much more readily distorted than properly base-paired DNA.
Explain how mismatch repair functions in E. coli.
MutS scans the DNA –> recognises a mismatch. The complex then recruits MutL. MutL activates MucH, an enzyme that causes an incision or nick on one of the strands. Nicking is followed by a helices (UvrD) and one of the three exonucleases. The helices unwinds the DNA and the exonucleases progressively digest the displaced single strand. The gap is filled in with DNA polymerase III.
How does the E. coli mismatch repair system know which of the two mismatched strands to repair?
E. coli tags the parental strands by transient hemimethylation. Dam methylase methylates a residues on both strands of the sequence 5’-GATC-3’. For a few minutes, until Dam methylase catches up the replication fork –> only the daughter DNA duplex will be methylated.
What are the proteins for mismatch repair called eukaryotic cells?
MutS –> MutS homologs (MSH)
MutL –> MutL homolog (MLH)
and PMS
How does the mismatch repair system know which of the two mismatched strands to repair in eukaryotes?
Okazaki fragments. MHS interacts with the sliding-clamp and would thereby be recruited tot he site of discontinuous DNA synthesis on the lagging strand.
What is the most common kind of hydrolytic damage?
Deamination of cytosine.
What occurs when cytosine is deaminated?
Cytosine undergoes spontaneous deamination, generating uracil. Uracil preferentially pairs with adenine and thus introduces that base in the opposite strand upon replication.
What occurs when adenine is deaminated?
Hypoxathine, which hydrogen bonds to cytosine rather than to thymine; guanine is converted to xanthine, which continues to pair with cytosine, although with only two hydrogen bonds.
Explain what occurs when DNA undergoes depurination.
By spontaneous hydrolysis of the N-glycosyl linkage, and this produces an basic site.
Explain what occurs when DNA undergoes alkylation?
Methyl and ethyl groups are transferred to reactive site on the base and phosphates int he DNA backbone.
Explain how DNA is damaged bu ultraviolet light?
Radiation with wavelength 260 nm is strongly absorbed by the bases, one consequence is the photochemical fusion of two pyrimidines that occupy adjacent positions not he same polynucleotide chain.
What does clastogenic mean?
Ionizing radiations and agents that cause DNA to break.
What are intercalating agents? What how do these agents causes short insertion and deletions?
Flat molecules containing several polycyclic rings that bind to the equally flat purine and pyrimidine bases of DNA. They slip between the bases in the template strand and causes the DNA polymerase to insert an extra nucleotide opposite the intercalated molecule.
What is excision repair?
The damaged nucleotide is note repaired but removed from the DNA. A new nucleotide is incorporated by DNA polymerase.
What is recombinational repair?
Repair when both strands are damaged. Sequence information is retrieved from a second undamaged copy of the chromosome.
What is photoreactivation?
Reverses the formation of pyrimidine dimers.
Explain base excision repair and nucleotide excision repair.
In base excision repair, an enzyme called glycosylase recognises and removes the damaged base by hydrolysing the glycosidic bond. The resulting basic sugar is removed from he DNA backbone in a further endonucleolytic step. Endonucleolytic cleavage also removes apurinic and apyrimidinic sugars that arise by spontaneous hydrolysis. (Endonuclease cuts the DNA at the 5’ position of the AP site, leaving the 3’ OH, exonuclease cuts the 3’position of the AP site, leaving the a 5’-phosphate. The resulting gap is fulled in by DNA polymerase I)
In excision repair, the enzyme recognises distortion to the shape of the helix. ATP hydrolysis promotes dimer formation by UvrA, which formed a complex with a dimer UvrB. This complex scans the DNA to identify distortions. UvrA leaves the complex, and the remaining UvrB dimer melts DNA locally around the distortion. UvrC forms a complex with UvrB and creates nicks 3’to the lesion and 5’ lesions. DNA helices UvrD releases the single-strand fragment from the duplex and DNA pol ! and ligase repair and seal the gap.
Transcription-coupled repair.
Rescuing RNA polymerase after being arrested by the presence of lesion in the transcribed strand of a gene.
Recruitment to the stalled RNA polymerase of nucleotides excision repair proteins.
When is NHEJ used?
When an unreplicated chromosome suffers a break and there is no sister chromosome present to serve as a template in the recombination-based DSB repair pathway.
Describe the mammalian pathway of NHEJ.
A heterodimer of Ku70 and Ku80 binds to the broken DNA and recruits the protein kinase DNA-PKcs. DNA PKcs in turn receipts Artemis, an enzyme having exonuclease and endonuclease activities, which processes the broken ends. Finally a complex of Ligase IV with XRCC4 and Cemunos-XLF joins the broken ends to each other.
Explain translesion synthesis.
Fail-safe mechanism that lets the replication machinery bypass the site of damage or tolerate the DNA damage.
Which family DNA polymerases catalyses translation synthesis?
In E. coli –> DNA Pol IV or DNA Pol V. DinB and UmuC are members of a distinct family of DNA polymerases found in many organisms known as the Y family of DNA polymerase.
