lecture 11 & 12- DNA mutations

Question 1

a mutation is a change in ___ that is propagated through ___

Answer 1

DNA sequence
cellular generations (has to be fixed mutation)

Question 2

when mutations occur in germ-line cells, they are ___

Answer 2

inheritable

Question 3

how can mutations be good

Answer 3

evolution
- some frequency of mutation is necessary to produce variability in which natural selection acts to drive evolution

Question 4

when does a DNA alteration become a stable, inheritable mutation?

Answer 4

only after the alternation is converted through replication into an incorrect base pair (such as an A-T pair where a G-C should be)

Question 5

what happens to the vast majority of damaged nucleotides that occur in mammalian cells?

Answer 5

fixed by DNA repair enzymes
- they can fix the alterations in DNA sequence, but if not fixed by next round of replication –> passed down (inheritable)

Question 6

point mutations are ___

Answer 6

single base pair substitutions

Question 7

transitions exchange a ___ for a ___
transversions exchange a ___ for a ___

Answer 7

purine for a purine (A–>G)
pyrimidine for a pyrimidine ( T–>C)

purine for a pyrimidine (A–>C)
pyrimidine for a purine (T–>G)

Question 8

name the two types of point mutations

Answer 8

transitions
transversions

Question 9

mutations of one or a few base pairs usually result from what?

Answer 9

errors in replication or damaged nucleotides

Question 10

what is more common- transitions or transversions?

Answer 10

transitions are 10 times more common

Question 11

a point mutation in the protein-coding region of a gene can result in what?
these point mutations can be classified as…

Answer 11

can result in an altered protein with partial or complete loss of function
- silent, missense, nonsense

Question 12

describe RNA complementarity to the DNA strands

Answer 12

RNA has same sequence as DNA top strand (coding- sense, nontemplate strand)
RNA is complementary to DNA bottom strand (noncoding, antisense, template strand)

Question 13

describe genetic code codons

Answer 13

DNA sequence encoding a protein is read in triplets (64 codons)
- 61 of 64 codons are sense codons for AA’s
- 3 of the 64 are nonsense- stop codons
UAA, UAG, UGA
- start codon = AUG (Met)
- code is degenerate (a single AA can be encoded by more than one codon)
. exception = Met (only encoded by AUG)

Question 14

name 2 rules in genetic code

Answer 14

1- no overlapping in triplets, read from a fixed point
. if it wasn’t, it would code different AA’s
2- open reading frame- a run of sense codons before a stop codon is encounters

Question 15

describe silent mutations

Answer 15

• mutations in non-coding or non-regulatory regions of DNA
• in coding regions- b/c of degeneracy, results in same AA
• ex: change from TTT to TTC is silent because both code for lysine (they are synonymous)

Question 16

describe missense mutations

Answer 16

any mutation within coding region leading to change in amino acid
- protein could have loss of function or gain of function if sequence change is in active site- severity depends on characteristics of the AA replaced (depends on nature and location of the change)
- ex: sickle-cell anemia- caused by missense mutation in gene that codes for beta subunit of hemoglobin - beta subunit has different shape, leads to aggregation of protein when with alpha subunit (change is glutamic acid –> valine)

Question 17

describe nonsense mutation

Answer 17

leads to stop codon, terminating translation
- generates a truncated protein without a complete AA sequence
- severity depends on location

Question 18

establishment of point mutations is a 2 step process…

Answer 18

1- create mutation- DNA polymerase incorporates an incorrect nucleotide
2- if mismatch is not repaired, it becomes a mutation in SECOND ROUND of replication

Question 19

insertions occur when…
deletions occur when…

Answer 19

insertions occur when one or more base pairs are added to the wild-type sequence
deletions occur with loss of one or more base pairs

Question 20

insertions and deletions may have no effect if…

Answer 20

not in genes or regulatory sequences

Question 21

an indel mutation of only one or two base pairs in the coding sequence of a protein throws off the reading frame after the mutation, resulting in a ___

Answer 21

frameshift mutation

Question 22

indels of 3 or multiples of nucleotides do what?

Answer 22

preserve the reading frame of the gene
- most common: insertion of 3 nucleotides (template slippage by DNA poly during replication) –> triplet expansion diseases, often CAG repeated triplet

Question 23

name 4 DNA alterations that lead to mutations

Answer 23

1- spontaneous hydrolysis
2- oxidative damage (ROS)
3- alkylation (alkylating agents)
4- radiation (UV, x-rays)

Question 24

spontaneous DNA damage by water can cause point mutations by ___ or ___

Answer 24

deamination of nitrogenous bases (C, G, or A) or loss of bases by hydrolysis

Question 25

deamination is the removal of a ___ group and changes ___

Answer 25

amino
changes identity of bases and their pairing properties

Question 26

describe deamination with cytosine

Answer 26

deamination of cytosine creates uracil
however, about 5% of cytosines in higher eukaryotes are methylated
- deamination of methylcytosine leads to thymine

Question 27

describe deamination with guanine or adenine

Answer 27

less frequent and produces abnormal bases
- guanine becomes xanthine, pairs with cytosine
- adenine becomes hypoxanthine, pairs with cytosine

Question 28

describe DNA damage by spontaneous hydrolysis

Answer 28

loss of bases by hydrolysis
- depurination much more frequent than depyrimidation
- 1 in 10,000 purines are lost from mammalian cell DNA every 24 hrs

Question 29

oxidative damage and alkylating agents can create ___ and ___

Answer 29

point mutations and strand breaks

Question 30

describe oxidative damage

Answer 30

sodium nitrate (common food preservative) and nitrosamines are converted to nitrous acid in the stomach (nitrous acid is potent mutagen)
- oxidative damage is possibly the most important source of mutagenic alterations in DNA
- ROS, H2O2, hydroxyl radicals (OH-) and superoxide radicals (O2-) arise during irradiation or as byproducts of aerobic metabolism
- oxdative damage includes modification of bases, sugar, removal of bases and strand breaks

Question 31

what is possibly the most important source of mutagenic alterations in DNA

Answer 31

oxidative damage

Question 32

oxidative damage includes…

Answer 32

modification of bases, sugar, removal of bases, and strand breaks

Question 33

describe oxidative guanine

Answer 33

oxidative guanine at position 8 is very mutagenic because it can pair with adenine, leading to a transversion if not repaired prior to replication
. instead of G-C, T-A –> one of the most common mutations in human cancers

Question 34

describe damage by alkylation

Answer 34

addition of an alkyl group (methyl, ethyl, propyl, isopropyl) to bases

Question 35

describe how guanine is damaged by alkylation

Answer 35

O6-methylguanine is very mutagenic because it tends to pair with T instead of C, leading to a transition (G–>A)

Question 36

give two examples of damage by alkylation

Answer 36

1- benzo pyrene (carcinogen, intercalating agent)- smoke of burning cigs, woodm coal
. in liver, becomes a reactive epoxide that can react with bases, especially guanine
2- nitrogen mustard gas- cross-linking agents that react with adjacent guanine residues, preventing replication and transcription

Question 37

how are DNA-damaging agents used in chemotherapy?

Answer 37

DNA-reactive agents used in chemo for cancer kill cells (cytotoxic) by creating broken chromosomes or stalled replication forks, both lead to cell death during cell division
- cross-linking agents
- intercalating agents- doxorubicin- stops replication forks, cells can’t divide anymore

Question 38

what is the Ames test used to identify

Answer 38

DNA-damaging, mutagenic chemicals in animals

Question 39

describe how the Ames test works

Answer 39

use histidine auxotrophic Salmonella typhimurium- mutation in biosynthetic pathway for histidine (requires histidine in growth medium)
- looks for his revertants
- when his- bacteria (S. typh) is plated in media without histidine and chemical-potential mutagen is added, cells show a reversion mutation (can synthesize his and grow in his-free medium) (the chemical led to mutations)

Question 40

most known human carcinogens result in what in the ames test?

Answer 40

increased mutation in the Ames test
- compounds identified as mutagens in an ames test require further testing to determine whether they are likely to be human carcinogens

Question 41

describe DNA damage caused by radiation

Answer 41

ionizing radiation: UV (in sunlight), cosmic rays, x-rays
- UV and other ionizing radiation: about 10% of all DNA damage caused by environmental agents
- UV can cause photochemical fusion of adjacent pyrimidines (pyrimidine dimers- covalent cross-links) – leads to no replication
- x-rays and gamma rays (higher energy): single or double-stranded DNA breaks

Question 42

UV light from the sun can cause ___ that stall ___

Answer 42

pyrimidine dimers that stall DNA polymerase during replication

Question 43

x rays and gamma rays cause ___

Answer 43

single and double stranded DNA breaks

Question 44

DNA-damaging agents can lead to mutations at ___ concentrations and can kill the cell at ___ concentrations

Answer 44

low
high