DNA polymerase minor groove interactions modulate mutagenic bypass of a templating 8-oxoguanine lesion.
A major base lesion resulting from oxidative stress is 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxoG) that has ambiguous coding potential. Error-free DNA synthesis involves 8-oxoG adopting an anti-conformation to base pair with cytosine whereas mutagenic bypass involves 8-oxoG adopting a syn-conformation to base pair with adenine. Left unrepaired the syn-8-oxoG/dAMP base pair results in a G-C to T-A transversion. During base excision repair of this mispair, DNA polymerase (pol) β is confronted with gap filling opposite 8-oxoG. To determine how pol β discriminates between anti- and syn-8-oxoG, we introduced a point mutation (R283K) to alter insertion specificity. Kinetic studies demonstrate that this substitution results in an increased fidelity opposite 8-oxoG. Structural studies with R283K pol β show that the binary DNA complex has 8-oxoG in equilibrium between anti- and syn-forms. Ternary complexes with incoming dCTP resemble the wild-type enzyme, with templating anti-8-oxoG base pairing with incoming cytosine. In contrast to wild-type pol β, the ternary complex of the R283K mutant with an incoming dATP-analogue and templating 8-oxoG resembles a G-A mismatched structure with 8-oxoG adopting an anti-conformation. These results demonstrate that the incoming nucleotide is unable to induce a syn-8-oxoG conformation without minor groove DNA polymerase interactions that influence templating (anti-/syn-equilibrium) of 8-oxoG while modulating fidelity.
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CHEMICAL CHEMICAL CHEMICAL CHEMICAL CHEMICAL CHEMICAL CHEMICAL CHEMICAL CHEMICAL CHEMICAL CHEMICAL CHEMICAL CHEMICAL CHEMICAL GENE-Y GENE-N GENE-N GENE-N GENE-Y GENE-Y GENE-N CHEMICAL GENE-N GENE-Y GENE-N CHEMICAL CHEMICAL CHEMICAL CHEMICAL CHEMICAL CHEMICAL CHEMICAL
8-oxoG adenine syn-8-oxoG dAMP 8-oxoG 8-oxo-7,8-dihydro-2'-deoxyguanosine anti- and syn-8-oxoG 8-oxoG 8-oxoG dCTP cytosine 8-oxoG anti-8-oxoG 8-oxoguanine pol β R283K minor groove DNA polymerase DNA polymerase (pol) β pol β R283K dATP R283K pol β DNA polymerase minor groove 8-oxoG 8-oxoG nucleotide syn-8-oxoG 8-oxoG 8-oxoG cytosine