Kinetic analysis of bypass of 7,8-dihydro-8-oxo-2'-deoxyguanosine by the catalytic core of yeast DNA polymerase ?.
Ontology highlight
ABSTRACT: Reactive oxygen species damage DNA bases to produce 7,8-dihydro-8-oxo-2'-deoxyguanosine (8-oxoG), which results in G:C to T:A transversions. To better understand mechanisms of dNTP incorporation opposite 8-oxoG, we performed pre-steady-state kinetic analysis of nucleotide incorporation using the catalytic core of yeast DNA polymerase ? (Pol ?core, residues 1-513) instead of full-length Pol ?, eliminating potential effects of the C-terminal C2H2 sequence motif on dNTP incorporation. Kinetic analysis showed that Pol ?core preferred to incorporate dCTP opposite 8-oxoG. A lack of a pre-steady-state kinetic burst for Pol ?core suggested that dCTP incorporation is slower than the dissociation of the polymerase from DNA. The extension products beyond the 8-oxoG were determined by LC-MS/MS and showed that 57% of the products corresponded to the correct incorporation (C) and 43% corresponded to dATP misincorporation. More dATP was incorporated opposite 8-oxoG with a mixture of dNTPs than predicted using only a single dNTP. The kinetic analysis of 8-oxoG bypass by yeast DNA Pol ?core provides further understanding of the mechanism of mutation at this oxidation lesion with yeast DNA polymerase ?.
SUBMITTER: Xue Q
PROVIDER: S-EPMC5193378 | biostudies-literature | 2016 Feb
REPOSITORIES: biostudies-literature
ACCESS DATA