Lecture 3 Flashcards
Point mutations definition
Mutations at a single point (most often a single base pair)
in a genome - SNPs (single nucleotide polymorphisms)
3 main categories of mutations
- point mutations
- substitution / indels (insertion or deletion of base-pairs)
- can also be deletion (loss) or large-scale rearrangements or insertion of mobile genetic elements
Many point mutations may occur____
Spontaneously i.e with no evident external cause
How do spontaneous mustions occur?
- require DNA synthesis
- can involve “mistakes” by DNA polymerase, the enzyme that synthesises DNA
- can arise through tautomerisation of bases - a base transiently flips into a different isomer that has different base-pairings properties (SNP result from Tautomerisation?)
Tautomerisation of bases affects ____-_____
Base-pairing
Example of normal base-pairs vs tautomeric base pairing
Base - changes structure - rearranges - giving cytosine a different form - imino form - this makes it much more chemically favourable to match to adenine
- same for thymine
How does the bacteria cut down the rate of spontaneous mutation
- DNA polymose ‘ checks’ each new base-pair
- if it has made a mistake it backs up and removes the incorrect base (over 99% effective)
- DNA synthesis then resumes
What serves as a prof reading function and how does it do this?
Both DNA pol 1 and DNA pol 111 serve a “proofreading” function by excising incorrectly inserted mismatched bases
- once the mismatched base is removed, the polymerase has another chance to add the correct complementary base
What is the importance of proof reading?
DNA proof-reading reduces the frequency of spontaneous mutation in bacteria
- in mice, lack of DNA proofreading causes increased mutations in mitochondrial DNA and pre mature aging of mice
Two ways of reducing spontaneous mutations
- proof reading by DNA pol 1 and 111
- the mismatch repair system
Properties of the mismatch repair system
- can detect non-matched “pairs” of bases in DNA molecules
- can determining which of the bases is wring
- can remove the wrong base
MMR (mismatch repair) process
- MutS protein detects mismatches of newly replicated DNA
- The binding of MutS to distortions in the DNA double helix recruits MutL and MuTH
- MutH cuts the newly synthesised strand containing the incorrect base
(- without the ability to discriminate between the parental and newly synthesised strands, the MMR system could not determine which base to excise - hoe does MutH know which is wrong? )
How does MMR (MutH) know which strand is newly synthesised (to fix)?
- Strand recognition by MutH is directed by adenine methylation at GATC sequences
- becuase adenine methylation occurs after DNA synthesis, newly synthesised DNA is temporarily unmodified, an this temporary absence of methylation directs repair to the new strand
DNA inside E.coli is chemically modified by methylation of adenine bases in the DNA - newly synthesised strands will not have a mathylated base - thus MutH can recognise it as the one to be fixed
Does not affect the pairing of the bases with thymines
What can the MMR system identify?
- system can identify mismatches AND
- distinguish the original (correct) and new (incorrect) strand of the DNA molecule
How MutS, MutH and Mut L repair in MMR
- MutS protein detects mismatches of newly replicated DNA
- The binding of MutS to distortions in the DNA double helix recruits MutL and MuTH
- Strand recognition by MutH is directed by adenine methylation at GATC sequences
- becuase adenine methylation occurs after DNA synthesis, newly synthesised DNA is temporarily unmodified, an this temporary absence of methylation directs repair to the new strand
- the MutH endonuclease cuts the unmethylated strand
- MutS - MutL also activates excision, which involves the DNA helicase UvrD and exonucleases
- DNA polymerase 3 repairs the single stranded gap, and DNA ligaments generates a continuous covalent DNA back bone
How mismatches arise form spontaneous chemical changes to bases
- Deamination, the hydrolytic removal of an amine group
E.g deamination converts cytosine to uracil - Uracil base pairs with adenine in replication, converting a C-G base pair into a T-A base pair
How do indel mutations arise?
Through an error during DNA replication
- DNA “slippage” results in loss or incorporation of a new base-pair into the DNA
Indels arise when loops in single stranded regions of DNA are stabilised by the “slipped mispairing” of repeated sequences in the course of DNA replication
Indel mutation - insertion diagram vs deletion diagram
- deletion occurs in parental strand
Rare base pairing - tautormisiration - why it martches better with the ‘wrong’ base
Hydrogen bonding?
How base tautomers cause mutaion
? Don’t get this?
