Deamination of Cytosine and Deamination of Cytosine and 5-methylcytosine5-methylcytosine

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Chemical MutagensChemical Mutagens

Intercalating AgentsIntercalating Agents EX, EX,

Ethidium Ethidium bromide, bromide, acridine acridine orangeorange

Can Can induce induce frameshiframeshift ft mutationmutationss

Uv Induced DimersUv Induced Dimers Thymine dimers Thymine dimers

and T-C dimersand T-C dimers Replication Replication

problemsproblems Interferes with the Interferes with the

ability of the T’s to ability of the T’s to base pair to the base pair to the opposite strand, opposite strand, and blocks DNA and blocks DNA replicationreplication

Other MutagensOther Mutagens Transposable elements—”jumping genes”.Transposable elements—”jumping genes”.

Major frameshift mutationsMajor frameshift mutations Factors in evolutionFactors in evolution

Mutator genes—mutations increase Mutator genes—mutations increase mutation rate. Four potent mutator genesmutation rate. Four potent mutator genes Mutant DNA pol III 3’Mutant DNA pol III 3’5’ exonuclease activity5’ exonuclease activity Mutant methylation enzymes(ex Mutant methylation enzymes(ex damdam)) Mutant enzymes in excision repair systemMutant enzymes in excision repair system Mutant enzymes in SOS systemMutant enzymes in SOS system

ReversionsReversions Mutations in an mutant can restore the Mutations in an mutant can restore the

wild type function (wild type function (reversion, back reversion, back mutation, or reverse mutationmutation, or reverse mutation))

Spontaneous or inducedSpontaneous or induced If mutation occurs at the site of the If mutation occurs at the site of the

original=original=True reversionTrue reversion Wild type restored by mutation at Wild type restored by mutation at

another site= another site= second site mutationsecond site mutation Second site in same gene= Second site in same gene= intragenic intragenic

suppressionsuppression Second site in another Second site in another gene=intergenic gene=intergenic

suppressionsuppression

Intragenic RevertantIntragenic Revertant TypesTypes

Same site Same site reversionreversion

Second site Second site revertant revertant

NOTE: shape NOTE: shape of R-groups of R-groups can also be a can also be a factor. EXfactor. EX

Reversion of Frameshift Reversion of Frameshift MutationsMutations

For reversion For reversion to be to be successfulsuccessful Reversion must Reversion must

be near original be near original site to reduce site to reduce # of aa altered# of aa altered

Section of Section of polypeptide polypeptide must be able to must be able to withstand withstand alteration alteration without without eliminating eliminating functionfunction

Intergenic SuppressionIntergenic Suppression Refers to a chnge in another gene Refers to a chnge in another gene

which suppresses or eliminates the which suppresses or eliminates the mutant phenotype. EXmutant phenotype. EX Multisubunit proteins—Mutation in one Multisubunit proteins—Mutation in one

subunit may be masked by mutation in subunit may be masked by mutation in another subunit (ex. restoring another subunit (ex. restoring hydrophobic patches)hydrophobic patches)

Suppression via suppressor tRNAsSuppression via suppressor tRNAs

Suppressor tRNASuppressor tRNA Nonsense mutation- aa codon Nonsense mutation- aa codon “Stop” “Stop”

Ex. AAGEx. AAGUAGUAG Some bacteria can “read through” these Some bacteria can “read through” these

mutations (though protein function may mutations (though protein function may be altered). HOW?be altered). HOW?

Mutant tRNA that has an anticodon that Mutant tRNA that has an anticodon that recognizes “Stop” as a reading codon.recognizes “Stop” as a reading codon. aa encoded depends on which tRNA is aa encoded depends on which tRNA is

mutatedmutated Not every suppressor restores normal Not every suppressor restores normal

functionfunction

Suppressor Mutants (cont’d)Suppressor Mutants (cont’d) EX. UUG (Leu)EX. UUG (Leu)UAG (Stop) (AUC anticodon)UAG (Stop) (AUC anticodon) A mutation in a tRNA resulting in “AUC” A mutation in a tRNA resulting in “AUC”

allows that tRNA to recognize “Stop”. allows that tRNA to recognize “Stop”. Can get suppression or partial Can get suppression or partial

suppressionsuppression NOTE: must be 2 copies of tRNA mutated. NOTE: must be 2 copies of tRNA mutated.

Why?Why? In any cell containing mutator, must In any cell containing mutator, must

also be a wild typealso be a wild type Suppressors allow survival, even if Suppressors allow survival, even if

sub-optimalsub-optimal

Termination of Translation in Termination of Translation in Suppressor StrainsSuppressor Strains

Problem: Must be a means of Problem: Must be a means of terminating translation.terminating translation.

HOW?HOW? Release factors still present, will Release factors still present, will

compete for the “Stop “ sitecompete for the “Stop “ site Many genes are double-Many genes are double-

terminatedterminatedEX. UAG-UAAEX. UAG-UAA

Photolyase

Fig. 20.39Fig. 20.40

O6-methylguanine methyl transferase

Repair by Direct Repair by Direct ReversalReversal

Excision Repair Excision Repair Most Most

common common repair repair mechanismmechanism

EX. Uvr EX. Uvr systemsystem

NOTE: NOTE: preferentiallpreferentially repairs y repairs dimers in dimers in essential essential regions of regions of genomegenome

UvRA recognizes UvRA recognizes damage and damage and binds w/UvrBbinds w/UvrB

UvRA released, UvRA released, UvrC bindsUvrC binds

UvrC nicks on UvrC nicks on both sides of both sides of damageUvrD damageUvrD unwinds regionunwinds region

Damaged strand Damaged strand releasedreleased

DAN pol IDAN pol I LigaseLigase

RecombinatioRecombinational Repairnal Repair

Sister Strand Sister Strand ExchangeExchange

Recombinational Repair, aka, Recombinational Repair, aka, Sister Strand ExchangeSister Strand Exchange

SOS Response SOS Response Is an inducible system of last resortIs an inducible system of last resort Also called error prone replication Also called error prone replication

because it inactivates the because it inactivates the proofreading function of DNA pol III. proofreading function of DNA pol III.

Turned on only when DNA damage is Turned on only when DNA damage is extremeextreme

Main players: recA and lexA and a Main players: recA and lexA and a battery of inducible enzymesbattery of inducible enzymes

SOS Response SOS Response