L7: DNA Mutability and Repair Flashcards
(55 cards)
1
Q
what are the 2 categories of mutations
A
- detrimental
- beneficial
2
Q
2 categories of mutations - detrimental
A
- more common
- most are deleterious
- cell cycle gene defects can lead to cancer
3
Q
2 categories of mutations - beneficial
A
genetic variation required for evolution
4
Q
what are the 2 consequences of mutations?
A
- regulatory or coding sequences of genes are altered and gets passed to progeny
- chromosomal structural changes that can then impede DNA replication and affect cell survival
5
Q
what are the 3 sources of mutations?
A
- DNA replication errors
- Chemical/environmental damage to DNA
- insertions generated by DNA elements (Transposons)
6
Q
DNA replication errors - cause
A
- nitrogenous base pairs are usually found in one tautomeric form
- instances of rare tautomer formation changes results in mispairing of bases
7
Q
DNA replication errors - tautomeric form of nitrogenous bases
A
- enol form: has a hydroxyl group
- keto: double bond on O
- usually C pairs with keto-G
- but if it changes to enol-G, it will fit in with T and DNA Pol cannot detect the mispair
8
Q
DNA replication errors - what are the classes of mutations
A
- transition
- transversion
- point mutations
- frameshift mutations
- chromosomal rearrangements
9
Q
DNA replication errors: classes - transitions
A
- same group different identity
- purine converted to purine (A ⇋ G)
- pyrimidine converted to pyrimidine (T ⇋ C)
10
Q
DNA replication errors: classes - transversion
A
- group is completely different
- purine converted to pyrimidine (A → T/C or G → C/T)
- pryrimadine converted to purine (T → G/A or C → G/A)
11
Q
DNA replication errors: classes - point mutation
A
- single base change
- has 3 types:
1. missense mutation
2. silent mutation
3. nonsense mutation
12
Q
DNA replication errors: point mutation - missense mutation
A
- changes the amino acid sequence of a protein
- damage depends on the type of amino acid being produced
13
Q
DNA replication errors: point mutation - silent mutation
A
- does not change the amino acid sequence of a protein
- least detrimental since it protein function does not change
14
Q
DNA replication errors: point mutation - nonsense mutation
A
- amino acid-specifying codon is changed to a stop
- most detrimental since its most likely to create a non-functional protein
15
Q
DNA replication errors: classes - frameshift
A
- mutation that alters the meaning of all downstream codons
- detrimental
16
Q
DNA replication errors: classes - chromosomal rearrangements
A
- larger in scale
- can result in additions or deletions of chromosomal regions
17
Q
DNA replication errors - chromosome-level mutations
A
- inversions
- translocation
- deletion
- duplication
18
Q
DNA replication errors: chromosome-level mutations - inversion
A
sections of a chromosome break and rotate before rejoining
19
Q
DNA replication errors: chromosome-level mutations - translocation
A
chromosome piece breaks and attaches to a different chromosome
20
Q
DNA replication errors: chromosome-level mutations - deletion
A
- segment of a chromosome is lost
- detrimental since it causes the loss in thousands of genes
21
Q
DNA replication errors: chromosome-level mutations - duplication
A
- additional copies of a chromosome segment is gained
- damage depends on molecular structure of the gene being effected
- genes that require a specific amount to be effective will make the mutation detrimental
22
Q
chromosome-level mutations: duplication - example of how it can be detrimental
A
- Charcot-Marie-Tooth (CMT) Disease
- caused by partial duplication of chr 17 including the PMP 22 gene (encodes myelin protein)
23
Q
DNA replication errors - DNA microsatellites
A
- sequences in DNA usually don’t code for genes (not always)
- repetition of 3 or 4 nucleotides over and over again
- hotspots for mutations and can cause diseases
24
Q
DNA replication errors: DNA microsatellites - why is it a hotspot for mutations?
A
- it can cause slippage of replication machinery
- some new DNA will slip and ‘bow out’
- replication will continue but it creates an extra repeat in the synthesized strand
- but can happen to the template, resulting in a loss of genetic material
25
DNA replication errors: DNA microsatellites - what's an example of a disease that can be caused by this?
- Huntington's Disease: degradation of nerve cells in the brain
- if errors create an increase in CAG repeats, individual will get an earlier onset of the disease
26
mismatch repair mechanism
- corrects mismatches incorporated due to DNA replication
- if mutation escapes proof-reading enzyme, fast-acting repair mechanisms must be in place to correct the error
27
mismatch repair mechanism - why must repair happen fast?
- during the first round of replication, if there is an error, a ‘bump’ is then created and detected (A-T vs A-G)
- if it escapes the proof-reading enzyme, the second round of replication will have the correct base pairing (A-T vs G-C)
- results in mutation no longer having bump and cannot be detected
28
mismatch repair mechanisms - *E. coli*
- uses **MutS**
- will then recruit **MutL** and **MutH**
29
mismatch repair mechanisms: *E. coli* - what is MutS?
- a mismatch repair protein that acts as a dimer
- it scans DNA and recognizes distortion due to mismatch
- binds to ATP and creates a kink in the duplex
- will then recruit **MutL** (endonuclease activity) and **MutH** (exonuclease activity - needs to terminal end to work)
30
mismatch repair mechanisms: *E. coli* - what is hapspens after MutS recruits MutL and MutH?
- they bind to the DNA and create a nick one one strand
- exonuclease (**MutH**) degredes DNA beyond mismatch (broad cut)
- DNA Pol III and DNA ligase fills in the space
31
mismatch repair mechanisms: *E. coli* - how does the cell know which strand to repair?
- enzyme **Dam methylase** methylates DNA
- it methylates A on both strands
- but immediately following replication, the DNA is hemi-methylated with only the parental strand haying methyl marks
- **MutH** only nicks the non-methylated strand (newly synthesized)
32
chemical/environmental damage to DNA - what are the types?
- hydrolysis
- radiation
33
chemical/environmental damage to DNA - hydrolysis
- interactions with water
- can cause deamination
34
chemical/environmental damage to DNA: hydrolysis - deamination of cystine
- water reacts with cystine and results in the loss of a NH2 group and creates uracil
- this makes the nucleotide convert to RNA form
- creates a G-A mutation (transition)
35
chemical/environmental damage to DNA: deamination of cystine - would it be worse if it was a thymine changing instead?
- no, consequences wouldn't be that bad
- bc if T changes to U, A will still fit inside it (T-A is the DNA equivalent to U-A in RNA)
36
chemical/environmental damage to DNA: hydrolysis - what does this answer?
- the question: why does DNA have T but RNA have U?
- possible answer: C → U mutation in DNA will make the cell know something is wrong since U is part of RNA
- if U was in DNA naturally, the cell wouldn't be able to see there is a mutation
37
chemical/environmental damage to DNA - radiation
1. UV light
2. γ-radiation and X-rays
38
chemical/environmental damage to DNA: radiation - UV light
- absorbed strongly by bases
- can result in chemical fusion of neighboring pyrimidines ("dimers") - also called thymine/pyrimidine dimers
- incapable of base-pairing and stall DNA synthesis
39
chemical/environmental damage to DNA: UV light - what are dimers?
- two pyrimidine structures near each other form covalent linkages between each other
- forms a cyclobutane ring and cannot interact with their partnered bases on the other strand
40
chemical/environmental damage to DNA: radiation - γ-radiation and X-rays
- cause double-stranded DNA breaks
- can be lethal to cell since intact chromosomes are required for DNA replication
41
chemical/environmental damage to DNA - how to repair broken DNA?
- direct reversal
- excision repair
- recombinal repair
- translesion DNA synthesis
42
chemical/environmental damage to DNA - direct reversal
- for Pyrimidine dimers (UV light)
- uses photoreactivation
43
chemical/environmental damage to DNA: direct reversal - photoreactivation
- enzyme **DNA photolyase** uses light energy to directly repair pyrimidine dimers
- does it by break the abnormal covalent bonds between the pyrimidines
44
chemical/environmental damage to DNA - excision repair
- two types:
1. base excision repair: replacement of specific, single base
2. nucleotide excision repair: stretch of nucleotides are replaced in DNA (more broad)
45
chemical/environmental damage to DNA: excision repair - base excision repair
- for small base modifications (deamination - hydrolytic damage)
1. enzyme **Glycosylase** hydrolyzes glycosidic bond of damaged base
2. **AP** ("apurinic" or "apyramidic") **endonuclease** removes abasic sugar
3. repair DNA Pol and DNA ligase restores correct nucleotide
46
chemical/environmental damage to DNA: excision repair - nucleotide excision repair
- for large DNA distortions (UV light)
- single stranded DNA segment containing lesion that distorts duplex (either side of DNA damage)
- enzymes in *E. coli*: UvrA-UvrB complex, UvrA, UvrB UvrC, UvrD
- higher eukaryotes (humans) have these as well, but different names
47
chemical/environmental damage to DNA: nucleotide excision repair - *E. coli*
1. **UvrA-UvrB complex** detects duplex distortion
2. **UvrA subunits** are released
3. **UvrB dimer** melts duplex
4. **UvrC** nicks either side of lesion
5. **UvrD helicase** unwinds and removes strad
48
chemical/environmental damage to DNA: nucleotide excision repair - repair enzyme mutation
- results in Xeroderma pigmentosum
- recessive disease caused by inability to repair UV-damaged DNA
- results in skin lesions and higher rates of skin cancer
49
chemical/environmental damage to DNA - recombinational repair
- used when both strands are damaged (radiation)
- done via non-homologous end joining (NHEJ)
50
chemical/environmental damage to DNA: non-homologous end joining (NHEJ) - how does it work?
- late resort and error-prone (mutagenic)
- DNA ends are processed for ligation (chewed off) by a variety of factors
- processing adds nucleotides and can introduce insertions or deletions
51
chemical/environmental damage to DNA - translesion DNA synthesis
- for UV damage
- last resort since its error-prone and mutagenic
- if lesion is not repaired and stalls DNA Pol, translesion synthesis can bypass the error altogether
52
chemical/environmental damage to DNA: translesion DNA synthesis - how does it bypass the error
- some parts of DNA are not synthesized (bc of dimer) division results in DNA break so chromosome is not replicated
- but one cell has too much DNA and another has too little
- the cell then employs a special class of translesion polymerases
53
chemical/environmental damage to DNA: translesion DNA synthesis - special class of translesion polymerases
- DNA Pol IV or V
- can add nucleotides independently of base-pairing
- not involved in everyday replication since they do not put in the correct nucleotide
- and DNA Pol III is not used since it cannot deal with this break
54
chemical/environmental damage to DNA: translesion DNA synthesis - how does the translesion polymerase access the DNA?
- the sliding clamp of DNA Pol III is ubiquinated
- this serves as a signal to recruit translesion polymerase which adds nonspecific nucleotides
- replicative DNA Pol can then resume synthesis
55
chemical/environmental damage to DNA: non-homologous end joining (NHEJ) and translesion DNA synthesis - why does are they used if they are error-prone
its better than not repairing the break since this can block replication or chromosomal loss resulting in death or tumor formation
