Lecture 11-DNA Recombination and Repair (Nakai) Flashcards
What is the primary purpose of homologous recombination?
to make repair possible!
Transition
replacing a purine with a purine or pyrimidine with a pyrimidine
transversion
replacing a purine with a pyrimidine or vice versa which results in either 2 single ring structures trying to bp or 2 double ring structures trying to bp
- CANNOT CANNOT result from a simple tautomeric shift
What are the main causes for DNA sequence changes? (3)
- cell physiology (O radicals, nucleases making nicks, cytoplasmic conditions causing damage to bases)
- Modification of bases from radiation/chemicals
- polymerase errors
Nitrous acid and hydoxylamine
- covalently modify bases leading to incorrect bping and h bonding
Why is T used instead of U in DNA?
probably to distinguish when DNA damage has been done because a very common mutation is the deamination of C –> U which happens every day in our cells despite whether or not a damaging agent is present
Why is U used in RNA?
its a common way to edit RNA. You can easily deaminate C –> U.
- the liver uses this to introduce premature stop codons and therefore truncated proteins
What is the most unstable nucleotide(s)?
- purines, especially guanine
What can potentially happen if guanosine has lost its guanine? (2)
- you can either introduce a mutation or cause a collapse at the replication fork.
What is the affect of alkylating a base? ex of a base prone to this?
it can change bping of the base
- ex: G
Name the 4 potential methylating/alkylating agents
- SAM
- Dimethylnitrosamine
- dimethylsulfate
- nitrogen mustard
What is the effect of radiation on bases?
Can shift equilibrium to favor higher energy tautomers (particularly A which will then bp with C)
Insertions/deletions are from
- movable genetic elements
Name the enzymes involved in BER in order
- DNA glycosylase
- AP endonuclease
- DNA Pol I + NTP
- ligase
Dam methylase
Recognizes GATC sequences on newly synthesized DNA strands that occur with statistical frequency and methylates the A!!!
MutS/MutL
- MutS recognizes damage in DNA before it has been methylated at its GATC sites by Dam methylase and binds to them using ATP
- MutL associates with MutS and will act as a molecular matchmaker
Explain the process of Mismatch repair
- Mut S finds DNA damage and binds (ATP required)
- MutL associates with MutS to act as a molecular matchmaker (ATP required)
- MutS/MutL complex search for nearest GATC site, recruit MutH (ATP required)
- MutH makes nick in the newly synthesized strand near the GATC site
- DNA Helicase II unwinds (ATP?)
- Depending on where the GATC sequence is relative to the error, certain exonucleases will remove a large segment of bps (ATP? either this and/or the helicase require ATP)
- DNA Pol III replaces the DNA since it is a LARGE GAP
- Ligase
- SSB involved
Lack of what correctional enzyme would result in a lethal mutation?
- uracil deglycosylase used in BER
Cause and effects of Hereditary Nonpolyposis Colorectal Cancer
- mutations in hMLH1 and hMSH2 and therefore ineffective mismatch repair
- cancers often seen in tissues that have rapid cell division: colon, upper urinary tract, stomach, small intestine, uterus, ovaries
Causes and effects of XP
- mutation in NER machinery leading to intolerance of sunlight bc of UV damage
- T dimers can’t be fixed and this either leads to cell death or likely cancer if the cell replicates through this
- few cells will replicate through this but some do and live to accumulate mutations
Cockaynes Syndrome and Trichothiodystrophy
- both are from a mutation within the NER as well
- mild UV sensitivity
- they lead to developmental issues such as growth retardation (Cockaynes) or mental retardation (Trich.)
Explain Transcription coupled repair
- when transcription machinery encounters lesion
- RNA Pol halts
- uncoupling proteins back up the pol without causing it to cease transcription while enzymes repair the lesion
- RNA Pol can then continue
- TFIIH involved
Mutations in TFIIH can result in ______
Cockaynes Syndrom or Trichothiodystrophy
Homologous recombination definition
exchange of DNA due to homology between sequences
Primary and secondary purpose of homolgous recombination
- Primary: restarting DNA replication forks when forks stall, role in meiosis
- Secondary: generation of genetic diversity–reassortment of maternal/paternal alleles
What role can homologous recombination play in meiosis?
tells which chromosomes need to be separated during meiosis
