Evidence for a Watson-Crick hydrogen bonding requirement in DNA synthesis by human DNA polymerase κ

William T. Wolfle, M. Todd Washington, Eric T. Kool, Thomas E. Spratt, Sandra A. Helquist, Louise Prakash, Satya Prakash

Research output: Contribution to journalArticlepeer-review

49 Scopus citations


The efficiency and fidelity of nucleotide incorporation by high-fidelity replicative DNA polymerases (Pols) are governed by the geometric constraints imposed upon the nascent base pair by the active site. Consequently, these polymerases can efficiently and accurately replicate through the template bases which are isosteric to natural DNA bases but which lack the ability to engage in Watson-Crick (W-C) hydrogen bonding. DNA synthesis by Polη, a low-fidelity polymerase able to replicate through DNA lesions, however, is inhibited in the presence of such an analog, suggesting a dependence of this polymerase upon W-C hydrogen bonding. Here we examine whether human Polκ, which differs from Polη in having a higher fidelity and which, unlike polη, is inhibited at inserting nucleotides opposite DNA lesions, shows less of a dependence upon W-C hydrogen bonding than does Polη. We find that an isosteric thymidine analog is replicated with low efficiency by Polκ, whereas a nucleobase analog lacking minor-groove H bonding potential is replicated with high efficiency. These observations suggest that both Polη and Polκ rely on W-C hydrogen bonding for localizing the nascent base pair in the active site for the polymerization reaction to occur, thus overcoming these enzymes' low geometric selectivity.

Original languageEnglish (US)
Pages (from-to)7137-7143
Number of pages7
JournalMolecular and cellular biology
Issue number16
StatePublished - Aug 2005

ASJC Scopus subject areas

  • Molecular Biology
  • Cell Biology


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