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PDBsum entry 3lzi
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Transferase/DNA
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PDB id
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3lzi
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References listed in PDB file
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Key reference
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Title
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Substitution of ala for tyr567 in rb69 DNA polymerase allows damp to be inserted opposite 7,8-Dihydro-8-Oxoguanine .
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Authors
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J.Beckman,
M.Wang,
G.Blaha,
J.Wang,
W.H.Konigsberg.
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Ref.
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Biochemistry, 2010,
49,
4116-4125.
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PubMed id
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Abstract
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Accurate copying of the genome by DNA polymerases is challenging due in part to
the continuous damage inflicted on DNA, which results from its contact with
reactive oxygen species (ROS), producing lesions such as
7,8-dihydro-8-oxoguanine (8-oxoG). The deleterious effects of 8-oxoG can be
attributed to its dual coding potential that leads to G --> T transversions.
The wild-type (wt) pol alpha family DNA polymerase from bacteriophage RB69
(RB69pol) prefers to insert dCMP as opposed to dAMP when situated opposite
8-oxoG by >2 orders of magnitude as demonstrated using pre-steady-state
kinetics (k(pol)/K(d,app)). In contrast, the Y567A mutant of RB69pol inserts
both dCMP and dAMP opposite 8-oxoG rapidly and with equal efficiency. We have
determined the structures of preinsertion complexes for the Y567A mutant with
dATP and dCTP opposite a templating 8-oxoG in a 13/18mer primer-template (P/T)
at resolutions of 2.3 and 2.1 A, respectively. Our structures show that the
8-oxoG residue is in the anti conformation when paired opposite dCTP, but it
flips to a syn conformation forming a Hoogstein base pair with an incoming dATP.
Although the Y567A substitution does not significantly change the volume of the
pocket occupied by anti-8-oxoG, it does provide residue G568 the flexibility to
move deeper into the minor groove of the P/T to accommodate, and stabilize,
syn-8-oxoG. These results support the hypothesis that it is the flexibility of
the nascent base pair binding pocket (NBP) in the Y567A mutant that allows
efficient insertion of dAMP opposite 8-oxoG.
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