PART III
What Genes Are and What They Do
CHAPTER
Anatomy and Function of a Gene: Dissection Through Mutation
OUTLINE
3.1 Mutations: Primary Tools of Genetic Analysis 3.2 What Mutations Tell Us About Gene Function 3.4 Examples found in eukaryotic gene regulation lecture
Types of mutations in the coding sequence of genes
Fig. 8.28a
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Mutations in the coding sequence of a gene can alter the gene product
Missense mutations replace one amino acid with another Conservative chemical properties of mutant amino acid are similar to the original amino acid
e.g. aspartic acid [(-)charged] glutamic acid [(-)charged]
Nonconservative chemical properties of mutant amino acid are different from original amino acid
e.g. aspartic acid [(-)charged] alanine (uncharged)
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Mutations in the coding sequence of a gene can alter the gene product (cont)
Nonsense mutations change codon that encodes an amino acid to a stop codon (UGA, UAG, or UAA)
Frameshift mutations result from insertion or deletion of nucleotides with the coding region
No frameshift if multiples of three are inserted or deleted
Silent mutations do not alter the amino acid sequence
Degenerate genetic code most amino acids have >1 codon
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A nonsense mutation in a protein-coding gene creates a truncated, nonfunctional protein
Fig 8.32a
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Nonsense suppression
A second, nonsense suppressing mutation in the anticodon of a tRNA gene allows production of a (mutant) full-length polypeptide
Fig 8.32b
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Mutations classified by their effect on DNA
Fig. 7.2a - c
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Mutations classified by their effect on DNA (cont)
Fig. 7.2d
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Mutations classified by their effect on DNA (cont)
Fig. 7.2e
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How natural processes can change the information stored in DNA
Depurination
1000/hr in every cell
Deamination of C C changed to U Normal C-G A-T after replication
Fig. 7.6a, b
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How natural processes can change the information stored in DNA (cont)
X-rays break the sugar phosphate backbone of DNA
Ultraviolet (UV) light causes adjacent thymines to form abnormal covalent bonds (thymine dimers)
Fig. 7.6c, d
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How natural processes can change the information stored in DNA (cont)
Irradiation causes formation of free radicals (e.g. reactive oxygen) that can alter individual bases 8-oxodG mispairs with A Normal G-C mutant T-A after replication
Fig. 7.6e
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Mistakes during DNA replication
Incorporation of incorrect bases by DNA polymerase is exceedingly rare (< 10-9 in bacteria and humans)
Two ways that replication machinery minimizes mistakes Proofreading function of DNA polymerase (Fig 7.7)
3'-to-5' exonuclease recognizes and excises mismatches
Methyl-directed mismatch repair (later in this chapter)
Corrects errors in newly replicated DNA
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DNA polymerases proofreading function
Mispaired base is recognized and excised by 3'-to-5' exonuclease of DNA polymerase
Improves fidelity of replication 100-fold
Fig. 7.7
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Rates of spontaneous mutation
Rates of recessive forward mutations at five coat color genes in mice
11 mutations per gene every 106 gametes
Mutation rates in other organisms
2 12 mutations per gene every 106 gametes
Fig. 7.3b
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Different genes, different mutation rates
Mutation rates are <10-9 to >10-3 per gene per gamete Differences in gene size Susceptibility of particular genes to various mutagenic mechanisms Average mutation rate in gamete-producing eukaryotes is higher than that of prokaryotes Many cell divisions take place between zygote formation and meiosis in germ cells
More chance to accumulate mutations
Can diploid organisms tolerate more mutations than haploid organisms?
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Unequal crossing-over can occur between homologous chromosomes
Pairing between homologs during meiosis can be out of register
Unequal crossing-over results in a deletion on one homolog and a duplication on the other homolog
Fig. 7.8a
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Transposable elements (TEs) move around the genome
TEs can "jump" into a gene and disrupt its function Two mechanisms of TE movement (transposition)
Fig. 7.8b
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How mutagens alter DNA: Chemical action of mutagen
Replace a base: Base analogs - chemical structure almost identical to normal base
Fig. 7.10a
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How mutagens alter DNA: Chemical action of mutagen
Alter base structure and properties: Hydroxylating agents add an OH group
Fig. 7.10b
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How mutagens alter DNA: Chemical action of mutagen (cont)
Alter base structure and properties (cont): Alkylating agents add ethyl or methyl groups
Fig. 7.10b
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How mutagens alter DNA: Chemical action of mutagen (cont)
Alter base structure and properties (cont): Deaminating agents remove amine (-NH2) groups
Fig. 7.10b
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How mutagens alter DNA: Chemical action of mutagen (cont.)
Insert between bases: Intercalating agents
Fig. 7.10c
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Mutations outside the coding sequence can disrupt gene expression
Fig. 8.28b
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Mutations classified by their effects on protein function
Table 8.2
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DNA repair mechanisms that are very accurate
Reversal of DNA base alterations Alkyltransferase removes alkyl groups
Photolyase splits covalent bond of thymine dimers
Homology-dependent repair of damaged bases or nucleotides
Base excision repair (Fig 7.11)
Nucleotide excision repair (Fig 7.12) Correction of DNA replication errors Methyl-directed mismatch repair (Fig 7.13)
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Base excision repair removes damaged bases
Different glycosylases cleave specific damaged bases Particularly important for removing uracil (created by cytosine deamination) from DNA
Fig. 7.11
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Nucleotide excision repair corrects damaged nucleotides
UvrA UvrB complex scans for distortions to double helix (e.g. thymine dimers) UvrB UvrC complex nicks the damaged DNA
4 nt to one side of damage 7 nt to the other side of damage
Fig. 7.12
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