Mutations
- somatic mutations: not transmitted from one generation to another
- germ-line mutations may be transmitted to 50% of offspring
Point mutations (base substitutions)
- silent/synonymous: no changes in aa sequence because of redundancy in genetic code
- missense/nonsynonymous: mutations causes 1 aa to be substituted for another, changing the aa sequence
- nonsense: an amino acid is converted to a stop codon
Indels
- cause frameshift s that alter reading frames, creating either nonsense or missense effects
- except when indels occur as multiples of 3 nucleotides
Classifying mutations by effect on functional phenotype
- a mutation that alters wildtype phenotype is a forwards mutation
- a reverse mutations changes mutant phenotype back to wildtype
- loss of funtion: protein function completely or partly lost (recessive inheritance)
- gain of function/radical: new gene product, or gene product in wrong tissue (dominant inheritance)
- neutral: missense mutation that results in non-significant change in protein function because one chemically similar aa is substituted for another or occurs in part of the protein not important to function
Transition vs transversions
- point mutations
- transitions occur when a purine is substituted by a purine, or a pyrimidine is substituted by a pyrimidine
- transversion: purine —> pyrimidine or vice versa
Reverse mutations
- a second mutation restores the function of polypeptide
- true reversions: one base change, and then change back
- suppressor mutations: where the first mutation is suppressed by a second mutation:
- intragenic suppressor (in the same gene)
- intergenic suppressor (in a different gene)
Intragenic suppressor mutations
- a missense mutation alters a single codon
- a second mutation at a different site in the same gene may restore the original amino acid
Eg. Original codon = UUA (Leu)
—> mutate to UUU (Phe)
—> 2nd mutation to CUU (Leu)
Intergenic suppressor mutation
- second mutation in another gene suppresses the effect of the first mutation
- eg. With nonsense mutation and rTNA has an anticodon for UAG so translation continues
How do mutations happen
- spontaneously (DNA machinery)
- induced by chemical and physical agents
Spontaneous mutations
- Tautomeric shifts (base tautomers) during DNA replication
- DNA strand slippage during DNA replication
- Misalignment of homologous chromosomes during crossing over (recombination) at meiosis 1
Tautomers of DNA bases
- usually a proton shift in the nucleotide
- changes the manner in which it bonds to opposite nucleotides
- A may bind with C
- T may bind with G
- base tautomers cause incorrect pairing during DNA replication
Slipped strand mispairing
- often causes indels
- common in sequences with repeats of the same base
- may cause the template strand to loop out or the synthesized strand to loop out
Misalignment during crossing over
- if homologous chromosomes misalign during crossing over, one product contains and insertion and one has a deletion
- usually because of repetitive sequences
Mutagens
-agents that cause mutation
- physical:
- ionizing radiation: cosmic, X-, gamma rays
- UV radiation from sunlight
- chemical:
- Base analogs
- Base modifying agents
- Intercalating agents
Ionization radiation
-cosmic rays: originate from the sun that are subatomic particles (99% simple protons like H and He nuclei)
- gamma rays: originally from radioactive decay
- produced by sub-atomic particles
- arise in astrophysical processes
- X-rays: emitted by electrons outside the atomic nucleus
- Lower energy gamma rays
- ionizing radiation creates free radicals which are highly reactive
- these can alter the structure of bases and break phosphodiester bonds in DNA
UV radiation
- electromagnetic radiation of lower energy than ionizing radiation
- from the sun
- can be generated by various types of lamps
- causes pyrimidine nucleotides next to each other to fuse together causing a kink in DNA strand
- can block replication or cause mutation
DNA repair enzymes used to correct damaged DNA
- nucleotide excision repair:
1. Protein recognized mismatches
2. Unwinds DNA in area of mismatch
3. Excises out nucleotides
4. Fills in correct nucleotides
Xeroderma pigmentosum
- autosomal recessive genetic disorder of DNA repair
- cant repair skin damage
- ability to repair mutations by UV light is deficient
Sterilization
-using UV light to sterilize equipment
Base analogs
-chemicals that appear similar to the normal bases in DNA, but cause incorrect base pairing and introduce point mutations during DNA replication
- eg. 5-Bromouracil very similar to thymine and cytosine
- normally pairs with adenine, but when ionized will base pair with guanine
-mutagenic effect occurs during DNA replication
Base modifying agents
-chemicals that modify groups on the normal bases in DNA that result in incorrect base-pairing and introduce point mutations during DNA replication
Intercalating agents
- chemicals that distort the normal stacking of bases in DNA resulting in insertion or deletion of a single base pair during replication
- flat molecules that insert between adjacent bases in DNA
- distort them by 0.68nm (the size of a base)
- first round of DNA replication, the DNA polymerase randomly selects any nucleoside triphosphate opposite the intercalating adjective
- result: frameshift mutations due to insertion of a base
Ames test
- assay for chemical mutagenicity
- measures the reversion of a mutant His- salmonella bacterial strain to His+ salmonella wild-type strain by potential mutagens
- His- salmonella cannot grow on minimal medium lacing essential aa histidine
- His+ salmonella will grow on minimal medium
- increased reversions of His- to His+ salmonella indicate the chemical is a mutagen, and a potential carcinogen
-inclusion of rat liver enzymes to mimic the chemical modification of potential mutagens in the human body
