Pre-Steady State Kinetic Studies of the Fidelity of Nucleotide Incorporation by Yeast DNA Polymerase δ
Pre-Steady State Kinetic Studies of the Fidelity of Nucleotide Incorporation by Yeast DNA Polymerase δ
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DOI：
10.1021/bi100556m
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发表时间：
2010-08-31
期刊：
影响因子：
2.9
通讯作者：
Washington, M. Todd
中科院分区：
文献类型：
--
作者：
Dieckman, Lynne M.;Johnson, Robert E.;Washington, M. Todd
Eukaryotic DNA polymerase delta (pol delta) is a member of the B family or polymerases and synthesizes most of the lagging strand during DNA replication. Yeast pol delta a is a heterotrimer comprised of three subunits: the catalytic subunit (Pol3) and two accessory subunits (Pol31 and Pol32). Although pol delta is one of the major eukaryotic replicative polymerase, the mechanism by which it incorporates nucleotides is unknown. Here we report both steady state and pre-steady state kinetic studies of the Fidelity of pol delta. We found that pol delta incorporates nucleotides with an error frequency of 10(-4) to 10(-5). Furthermore, we showed that for correct versus incorrect nucleotide incorporation, there arc significant differences between both pre-steady state kinetic parameters (apparent K-d(dNTP) and k(pol)). Somewhat surprisingly, we found that pol delta synthesizes DNA at a slow rate with a k(pol) of similar to 1 s(-1). We suggest that, unlike its prokaryotic counterparts, pol delta requires replication accessory factors like proliferating cell nuclear antigen to achieve rapid rates of nucleotide incorporation.
