Lecture 5 - Mutation Flashcards
What is the keto-enol equilibrium?
The DNA bases exist in these two tautomers - isomers of each other - same chemical formula but different chemical arrangement of the atoms
Keto form - hydrogen on one end, double bonded oxygen on other end. Hydrogen on other end of molecule to oxygen will drift along molecule and attach to oxygen in the enol form, freeing off the relatively polar nitrogen to make new hyderogen bonds.
The keto form is strongly favoured by the equilibrium
What happens when tautomers mispair?
If the T changes to enol form three hydrogen bonds are formed between the bases rather than two so would pair with G rather than A.
Same principle applies for G-C base pairs - amino-imino tautomers
If you have change into enol form of T, then the DNA replication fork passes through, then the G nucleotide will get erroneously paired with the T enol. Either this is caught and repaired by replication machinery or it will convert back to its keto form in next round of replication and pair correctly, but the changed G will now pair with C and this is where we have a mutation.
Describe how base slipping causes indels
DNA backbone has a degree of “give” when the replication fork goes through. Little loops form and when the polymerase passes through the template strand is different length to the new strand
If the slippage happens in the new strand, there is +1 addition, if in template strand, -1 deletion
Indels - insertions/deletions, tandem repeats - repeated short sequences of base pairs
Polymerase will slide along these repeats and might skip along one (deletion) or skip back one (addition)
What are spontaneous lesions?
Happens a lot in humans. Backbone is there and is continued but base pair is missing
missing information on template strand, leads to random incorporation of bases, frequently A
What is spontaneous demamination?
c deaminated to U
What is the effect of the oxidation of bases?
Another way of damaging bases is oxidation, e.g oxidation of guanine. The result is an additional hydrogen bond donor and that guanine can now base pair with A (instead of C). Very frequent, reactive oxygen species is generated through normal metabolism and at higher levels in situations of cellular stress and irradiation of cells. Radiation=more oxidation = more mutation
What is the Ames test?
Ames (named from Bruce Ames) tests takes into account this fact that some compounds need to be metabolised. So add your aflatoxin or benzopyrine into the liver extract, then put onto a plate of salmonella bacteria mutant for genes in the histidine synthesis pathway and grow on His- media. If you put your paper disc on and you get colony growth this indicates that you have revertants
