Lecture 5- DNA damage and mutation

Question 1

Mutation

Answer 1

An alteration to the genetic material (DNA or RNA) that produces a heritable change in the nucleotide sequence

Question 2

Wild type

Answer 2

The standard form of a gene or an organism

Question 3

Mutant

Answer 3

The altered gene or organism produced by mutation

Question 4

Forward mutation

Answer 4

A process that converts a wild type to a mutant

Question 5

Reversion

Answer 5

A process that converts a mutation to a wild type

Question 6

Phenotype

Answer 6

A description of what an organism looks like

Question 7

Genotype

Answer 7

A description of the genes of an organism

Question 8

Mutagen

Answer 8

A chemical or physical agent that causes mutation

Question 9

Spontaneous mutation

Answer 9

Genetic changes that arise naturally during the life of an organism

Question 10

Induced mutation

Answer 10

Genetic changes caused by a specific mutagen

Question 11

What are the 3 different types of mutation?

Answer 11

Global change
Localised change
Base pair substitution

Question 12

Global change mutation

Answer 12

Large scale genomic changes
Chromosomal aberrations: deletions, insertions, duplications and inversions
Genome re-arrangement: redistribution if genetic material between chromosomes (translocation) that often arise from chromosome breakage
Change in chromosome number: trisomy of chromosome 21 (Down’s syndrome) which usually arises from mistakes in chromosome segragation at cell division

Question 13

Localised change

Answer 13

Affects a small number of nucleotides
Base substitution: point mutation (single base, frame shifts)
Delection, insertion
Duplication: sequence is repeated
Inversion: sequence inverted
Translocation or transposition: movement of a piece of DNA from one location to another

Question 14

Base pair substitutions

Answer 14

Point mutation: a mutation that results in the substitution of one base pair for a different base pair

These can be 2 types:
Transition mutation: change a purine for a purine or change a pyrimidine for a pyrimidine
Transversion mutation: change a purine for a pyrimidine or change a pyrimidine for a puring

Question 15

What are the 5 possible outcomes of mutations?

Answer 15

Silent/samesense: no effect
Missense: amino acid substitution
Nonsense: amino acid changed to stop codon
Readthrough: stop codon changed to amino acid
Frame shift: base pair deletions or insertions in a gene induce a shift in the reading frame

Question 16

What are the 4 different effects of point mutation?

Answer 16

No effect: silent
Missense: protein polymorphism
Change of function
Loss of function

Question 17

Why are adduct and lesion interchangeable?

Answer 17

They both mean DNA damage

Question 18

What are the 2 main categories of damage?

Answer 18

Affecting a single strand of DNA

Affecting both strands of DNA

Question 19

DNA damage that affects a single strand of DNA

Answer 19

Easier to fix as there is still an undamaged strand that can acts as a template
Adduct/lesion: 1 or 2 nucleotides altered e.g. methylated base
Nick
Mismatch

Question 20

DNA damage that affects both strands of DNA

Answer 20

Replicated adduct: a gap opposite the adduct as DNA polymerase skips the damaged region
Chromosomal break/interstrand crosslink

Question 21

What is a spontaneous mutation?

Answer 21

Damage from spontaneous degradation of DNA (endogenous factors)
Loss of bases
Loss of amine groups from bases
Mutations from damage by metabolic products (reactive oxygen species)
Fixation of mismatches and other mistakes by DNA polymerases

Question 22

What are the 3 different types of spontaneous mutation?

Answer 22

Depurination
Deamination
Tautomeric shift

Question 23

Depurination (spontaneous mutation)

Answer 23

The glycosidic bond between the base and sugar is cleaved by hydrolysis resulting in an apurinic or apyrimindic site/abasic site as a base has dropped out. Loss of adenine or guanine are the most common. A human cell loses several thousand purines each day.

Question 24

Deamination (spontaneous mutation)

Answer 24

Amine groups on the rings of bases are susceptible to spontaneous oxidation to aldehyde groups- deamination. This alters the pairing properties of the bases e.g. cytosine deaminates to uracil which can base pair with adenine when replication occurs

Question 25

Tautomeric shifts (spontaneous mutation)

Answer 25

Bases in DNA can occur in several forms, tautomers, which differ in the positions of their atoms and in the bonds between the atoms. The keto form of each base is normally present in DNA, whereas the imino and enol forms of bases are rare. DNA bases can isomerise with the different isomers having different base pairing properties. These changes are a significant source of spontaneous mutation.

Question 26

Tautomeric shift of guanine

Answer 26

Guanine will undergo a tautomeric shift to its rare enol form G* prior to replication. In its enol form it will pair with thymine and post replication is reverts back to its normal keto form.

Question 27

Describe tautomeric shifts, step by step

Answer 27

Hydrogen atoms move to form bases with altered hydrogen bonding properties creating a tautomeric shift. When a tautomeric shift occurs in adenine, adenine can bind to cytosine. A tautomeric shift in guanine allows it to bind to thymine. This error is passed on through replication and onto mRNA which can lead to a different amino acid being incorporated into a protein.

Question 28

What is an induced mutation?

Answer 28

``` damdage from external (Exogenous) sources. Mis-repair of damage caused by: Radiation Alkylation of bases Crosslinking agents Intercalated molecules ```

Question 29

What are the 3 main causes of induced mutations?

Answer 29

Exogenous oxidative damage Radiation Chemical mutations

Question 30

Endogenous oxidative damage - example

Answer 30

The products of oxidative processes that form the basis of aerobic life. E.g. thymine glycol formed by hydroxyl radical attach of thymine, thymine glycol blocks DNA replication

Question 31

Damage limitation of endogenous oxidative damage

Answer 31

Oxidation of FADH2 on flavoproteins leads to production of hydrogen peroxide and superoxide. Catalases (Kats), peroxidases (e.g. alkyl hydroperoxide reductase, Ahp) and superoxide dismutases (SODs) minimise the accumulation of these two oxidants. Glutathione is a tripeptide used as a cellular antioxidant to mop up ROS (reactive oxygen species)

Question 32

What are the indirect effects of radiation induced mutations?

Answer 32

Particle interacts with other molecules which then interact with the DNA e.g. radiolysis of water to give hydroxyl radicals which damage bases

Question 33

What are the direct effects of radiation induced mutations?

Answer 33

Particle imparts its energy directly to the DNA molecule. Breakage of the bonds that hold the sugar phosphate backbone together. Ultraviolet light has a direct effect on DNA.

Question 34

What are the main products of UV irradiation- radiation induced mutations

Answer 34

Cyclobutane pyrimidine dimers and 6-4 photoproducts Formed at adjacent pyrimidines resulting in covalently fused dimers. Lesions interfere with normal base pairing and can cause base substitutions. Double helix is distorted- this damage is bulky lesion.

Question 35

What is the order of lesions that are formed preferentially - radiation induced mutations

Answer 35

TT TC CT CC

Question 36

6-4 photoproduct - radiation induced mutation

Answer 36

The C6 carbon of one pyrimidine covalently links with the C4 carbpn of an adjacent pyrimidine. The backbone becomes distorted resulting in a bulky leasion

Question 37

Chemical mutagens, alkylating agents - induced mutation

Answer 37

Natural and artificial agents Alkylating agents: add methyl or ethyl groups to bases. This can alter the pairing properties and are called miscoding lesions. Some block replication.

Question 38

Chemical mutagens, Benz(o)pyrene - induced mutation

Answer 38

Carcinogenic ingredient of hydrocarbons in chimney soot, cigarette smoke and car exhausts if metabolised to an epoxide that acts on the N2 group of guanine.

Question 39

Chemical mutagens, Mutagens produced by metabolic activation - induced mutation

Answer 39

Many mutagens are not active and require bioactivation to electrophiles that bind covalently to DNA. Acetylaminofluorene is an aromatic amine originally used as an insecticide that is converted to an alkylating agent by esterification.

Question 40

Chemical mutagens, DNA crosslinking - induced mutation

Answer 40

Certain chemicals covalently join two bases in complementary strands to form an interstrand crosslink. DNA crosslinks prevent separation of two strands so block transcription and DNA replication. Interstrand crosslinks are typically produced by bifunctional agents

Question 41

Chemical mutagens, DNA intercalating agents - induced mutation

Answer 41

DNA double helix is partly held together by stacking interactions. Some chemicals are capable of intercalating the stacked DNA bases. Intercalation can cause single nucleotide insertions and deletions blocking replication/transcription.