Human DNA polymerase κ bypasses and extends beyond thymine glycols during translesion synthesis in vitro, preferentially incorporating correct nucleotides

Paula L. Fischhaber, Valerie L. Gerlach, William J. Feaver, Zafer Hatahet, Susan S. Wallace, Errol C. Friedberg

Research output: Contribution to journalArticle

91 Scopus citations

Abstract

Human polymerase κ (polκ), the product of the human POLK (DINB1) gene, is a member of the Y superfamily of DNA polymerases that support replicative bypass of chemically modified DNA bases (Ohmori, H., Friedberg, E. C., Fuchs, R. P., Goodman, M. F., Hanaoka, F., Hinkle, D., Kunkel, T. A., Lawrence, C. W., Livneh, Z., Nohmi, T., Prakash, L., Prakash, S., Todo, T., Walker, G. C., Wang, Z., and Woodgate, R. (2001) Mol. Cell 8, 7-8; Gerlach, V. L., Aravind, L., Gotway, G., Schultz, R. A., Koonin, E. V., and Friedberg, E. C. (1999) Proc. Natl. Acad. Sci. U. S. A. 96, 11922-11927). Polκ is shown here to bypass 5,6-dihydro-5,6-dihydroxythymine (thymine glycol) generated in two different DNA substrate preparations. Polκ inserts the correct base adenine opposite thymine glycol in preference to the other three bases. Additionally, the enzyme correctly extends beyond the site of the thymine glycol lesion when presented with adenine opposite thymine glycol at the primer terminus. However, steady state kinetic analysis of nucleotides incorporated opposite thymine glycol demonstrates different misincorporation rates for guanine with each of the two DNA substrates. The two substrates differ only in the relative proportions of thymine glycol stereoisomers, suggesting that polκ distinguishes among stereoisomers and exhibits reduced discrimination between purines when incorporating a base opposite a 5R thymine glycol stereoisomer. When extending beyond the site of the lesion, the misincorporation rate of polκ for each of the three incorrect nucleotides (adenine, guanine, and thymine) is dramatically increased. Our findings suggest a role for polκ in both nonmutagenic and mutagenic bypass of oxidative damage.

Original languageEnglish (US)
Pages (from-to)37604-37611
Number of pages8
JournalJournal of Biological Chemistry
Volume277
Issue number40
DOIs
Publication statusPublished - Oct 4 2002

    Fingerprint

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this