2oyq
From Proteopedia
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Crystal structure of RB69 gp43 in complex with DNA with 5-NIMP opposite an abasic site analog
Overview
Damage to DNA involving excision of the nucleobase at the N-glycosidic, bond forms abasic sites. If a nucleotide becomes incorporated opposite an, unrepaired abasic site during DNA synthesis, most B family polymerases, obey the A-rule and preferentially incorporate dAMP without instruction, from the template. In addition to being potentially mutagenic, abasic, sites provide strong blocks to DNA synthesis. A previous crystal structure, of an exonuclease deficient variant of the replicative B family DNA, polymerase from bacteriophage RB69 (RB69 gp43 exo-) illustrated these, properties, showing that the polymerase failed to translocate the DNA, following insertion of dAMP opposite an abasic site. We examine four new, structures depicting several steps of translesion DNA synthesis by RB69, gp43 exo-, employing a non-natural purine triphosphate analogue, 5-nitro-1-indolyl-2'-deoxyriboside-5'-triphosphate (5-NITP), that is, incorporated more efficiently than dAMP opposite abasic sites. Our, structures indicate that a dipole-induced dipole stacking interaction, between the 5-nitro group and base 3' to the templating lesion explains, the enhanced kinetics of 5-NITP. As with dAMP, the DNA fails to, translocate following insertion of 5-NIMP, although distortions at the, nascent primer terminus contribute less than previously thought in, inducing the stall, given that 5-NIMP preserves relatively undistorted, geometry at the insertion site following phosphoryl transfer. An open, ternary configuration, novel in B family polymerases, reveals an initial, template independent binding of 5-NITP adjacent to the active site of the, open polymerase, suggesting that closure of the fingers domain shuttles, the nucleotide to the active site while testing the substrate against the, template.
About this Structure
2OYQ is a Protein complex structure of sequences from Enterobacteria phage rb69 with and as ligands. Active as DNA-directed DNA polymerase, with EC number 2.7.7.7 Full crystallographic information is available from OCA.
Reference
Caught bending the A-rule: crystal structures of translesion DNA synthesis with a non-natural nucleotide., Zahn KE, Belrhali H, Wallace SS, Doublie S, Biochemistry. 2007 Sep 18;46(37):10551-61. Epub 2007 Aug 24. PMID:17718515
Page seeded by OCA on Wed Jan 23 12:43:20 2008
