2c22: Difference between revisions

DNA polymerases insert dATP opposite the oxidative damage product, 7,8-dihydro-8-oxodeoxyguanosine (8-oxoG) instead of dCTP, to the extent of, >90% with some polymerases. Steady-state kinetics with the Y-family, Sulfolobus solfataricus DNA polymerase IV (Dpo4) showed 90-fold higher, incorporation efficiency of dCTP > dATP opposite 8-oxoG and 4-fold higher, efficiency of extension beyond an 8-oxoG:C pair than an 8-oxoG:A pair. The, catalytic efficiency for these events (with dCTP or C) was similar for G, and 8-oxoG templates. Mass spectral analysis of extended DNA primers, showed >/=95% incorporation of dCTP > dATP opposite 8-oxoG., Pre-steady-state kinetics showed faster rates of dCTP incorporation, opposite 8-oxoG than G. The measured K(d)(,dCTP) was 15-fold lower for an, oligonucleotide containing 8-oxoG than with G. Extension beyond an, 8-oxoG:C pair was similar to G:C and faster than for an 8-oxoG:A pair, in, contrast to other polymerases. The E(a) for dCTP insertion opposite 8-oxoG, was lower than for opposite G. Crystal structures of Dpo4 complexes with, oligonucleotides were solved with C, A, and G nucleoside triphosphates, placed opposite 8-oxoG. With ddCTP, dCTP, and dATP the phosphodiester, bonds were formed even in the presence of Ca(2+). The 8-oxoG:C pair showed, classic Watson-Crick geometry; the 8-oxoG:A pair was in the syn:anti, configuration, with the A hybridized in a Hoogsteen pair with 8-oxoG. With, dGTP placed opposite 8-oxoG, pairing was not to the 8-oxoG but to the 5' C, (and in classic Watson-Crick geometry), consistent with the low frequency, of this frameshift event observed in the catalytic assays.

2C22 is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Sulfolobus_solfataricus Sulfolobus solfataricus] with CA and DGT as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/DNA-directed_DNA_polymerase DNA-directed DNA polymerase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=2.7.7.7 2.7.7.7] Structure known Active Site: AC1. Full crystallographic information is available from [http://ispc.weizmann.ac.il/oca-bin/ocashort?id=2C22 OCA].  

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