Mutations Flashcards
Is DNA replication 100% accurate? What is its error rate if it isn’t?
• DNA replication is accurate, but not 100% accurate
o DNA replication has a template, which is why it is precise
o Reasonably high error rate-10-9 is error rate of incorporating new mutation through each DNA replication cycle
Is genetic variation per generation predictable or random? Include an example
• Predictable rate of genetic variation per generation
o Incorporated through DNA replication and other mutation events
o Humans have about 70 new base pairs different from parents’ genomes due to changes in germline through mutation
Describe Darwin’s variation in finch beaks, what he obesrved from that and what gene is responsible for this variation
• Darwin- variation in finch beaks
o Some finches with blunt beaks
o Some finches with sharp pointed beaks
o Inferred that there was a difference in way finches developed their beaks
o Direct relationship to physical changes and DNA sequence
Gene responsible for beak shape is ALX1 (a gene in region D expressed in craniofacial region)
• In mammals with ALX1 gene, mutation causes craniofacial defects
Are bases chemically inert (cannot change)?
• The bases can change in chemical forms
What two forms can bases have and which is more common/rarer and why?
• The bases can change in chemical forms
o Switch between keto and enol form
Normal -keto form which are more energetically stable than enol forms
Rare- enol form called tautomers
What are tautomers?
Enol forms of nucleotide bases
How are enol forms different from base keto forms?
o Different types of hydrogen bonds form between bases when they’re in their enol form vs their keto form
Tautomeric forms change position of the double bonds and proton shifts within structure of the bases, forming less stable pairings
o Normal keto (common) form: makes TA (2 hydrogen bonds) and CG (3 hydrogen bonds) base pairing (Watson-Ccrick base pairs)
Describe how mispairings in DNA replication relating to different base forms is possible and what base pairs this result in
• This can lead to mispairings in DNA replication-Tautomeric shift of nucleotide bases can cause mispairings of bases and can result in newly synthesised DNA pair having a mutation
o When have Cytosine in enol form and adenine in common form, these two bases can base pair with 2 hydrogen bonds (slightly energy stable, but not as much as TA pair)
o When have thymine in common form and guanine in rare form, these two bases can base pair with 3 hydrogen bonds
Describe how frequently enol-keto mispairing occurs and what can be done to reduce this
o There is mispairing 1 in 100,000 bases -> tens of thousands of errors in 1 replication of mammalian genome
100 fold reduction of these mutations by error correction in DNA replication
Describe the process of mispairing in DNA replication and why it is a problem
• Process of DNA replication-tautomers
o Standard nucleotides and base pairing
o Tautomeric shift of nucleotide from keto form to enol form
o Replication occurs
o The enol tautomer mispairs with a keto form nucleotide that it normally doesn’t pair with
o Strands separated
o Tautomeric shift of enol form back to keto form
o Replication occurs
o There is a mutation in the newly synthesised strand in second step of replication-mutant strand of DNA in ½ the progeny
Where do most mutations take place?
In DNA replication
What is the mechanism of repairing mispaired bases during replication? Is this mechanism slow or fast and why?
• Polymerase exonuclease
o Polymerase removes most mispaired bases during replication by measuring distance between nucleotides (mispaired nucleotides have different distances between each other than normal ones)
o Removes wrong ones via 3’ exonuclease activity
o New opportunity for pairing and inserts right base pairs
o Proofreading slows down replication but makes it more accurate
o Fast replication done by error-prone polymerases with no exonuclease function so make it less accurate (sometimes these are used in PCR)
Describe the purpose of base excision repair and its mechanism
• Base excision repair
o Breakage of bond connecting the base to the sugar-phosphate backbone
o Loss of base creates an apurinic or apyrimidinic site (AP)
o AP is recognised by AP endonuclease, which cleaves the backbone on each side of the base
o DNA polymerases can fill in this gap
How was the first bacterial repair system identified by?
• First bacterial repair system identified by:
o Phenotype- high rate of mutation in E.Coli
o Genotype- mut genes were mutant
Mut genes wild type- DNA repair is normal
Mut genes are mutant- DNA repair is less efficient
Describe the purpose of mismatch repair and how it occurs
o Catches mismatches missed by DNA replication proofreading or that have occurred at another time
o Mismatch is detected with proteins
o The mismatch repair proteins remove the mispaired base and surrounding bases on the new strand
Removes about 10 nucleotides or so around the mismatch and does it on newly synthesised strand
o DNA polymerase fills the gap and ligase seals the nicks
What is the difference in the detection step in mismatch repair between bacteria and other organisms?
In bacteria, template strand is more highly modified than newly synthesised strand which is how it knows what to use as a template. Doesn’t happen in all organisms
• Old strand- has methylated adenines
• New strand- unmethylated adenines
Describe how direct repair occurs
• Direct repair
o Many environmental agents attach methyl groups to bases, especially guanine which can cause guanine to mispair with thymine
o Chemically modified bases can be directly repaired by removal of methyl groups by methyltransferases
Describe how break repair occurs (purpose and process)
• Break repair
o Double-stranded breaks to DNA are repaired by reattaching the broken end to another DNA molecule
o Two different processes, with many variations, are used
o Commonly, the DNA molecules are not similar in sequence, so the breaks are repaired by non-homologous end joining
Less commonly, the DNA molecules are highly similar in sequence, and homologous recombination occurs
What is the danger of UV light?
• UV light causes lots of mutations
Describe the mutation induced by UV light and how it triggers repair
• UV light has high energy and causes pyrimidine dimer
o Mutation event that isn’t normal-physically joins two adjacent pyrimidines (usually Ts)
• This leads to a bulge in the helix which triggers repair
What is photoreactivation and in what organisms does it occur?
• Photoreactivation- Bacteria, single-celled eukaryotes, plants and some animals (not humans)
o Pyrimidine dimers repaired by photoreactivation
o Photolyase binds the thymine dimer and enzyme photolyase uses energy from visible light to break the bonds between pyrimidine dimers
o Photolyase is encoded by the E.coli phr (photoreactive repair) gene
Describe how UV repair proceeds in humans
• UV repair in humans-
o Nucleotide excision repair
Changes to multiple adjacent bases or more extensive damage
When damage to the nucleotides is greater than a single base, the damaged region is recognised: the strands are separated, and oen strand is degraded to create a gap of 10-20 nucleotides
This gap is filled in by DNA polymerases
While the overall process is similar in bacteria and eukaryotes, the proteins are not highly conserved
What is the consequence of people who have defects in process of nucleotide excision repair?
o People who have defects in process of nucleotide excision repair are affected by xeroderma pigmentosa and are extremely sensitive to UV light, often have basal cell cacinomas and have reduced life expectancy
People have mutations in 1 of 7 genes of encoding repair proteins
What are 3 different types of mutations?
• Indels
o Small insertions or deletions (generally 1,2,3 or 4 base pairs)
• Transitions
o A purine (A,G) is replaced by a purine or a pyrimidine (C,T) is replaced by a pyrimidine
• Transversions
o A purine is replaced by a pyrimidine or vice versa