Enhanced ribonucleotide incorporation by an O-helix mutant of Thermus aquaticus DNA polymerase I

M. Ogawa, A. Tosaka, Y. Ito, S. Yoshida, M. Suzuki

研究成果: Article査読

13 被引用数 (Scopus)

抄録

The O-helix of DNA polymerases has been implicated in substrate discrimination and replication fidelity. In this study, wild-type Thermus aquaticus DNA polymerase I (Taq pol I) and an O-helix mutant A661E was examined for their ability to discriminate between ribonucleotides and deoxyribonucleotides. Steady-state nucleotide extension kinetics were carried out using a template cytidine and each nucleotide dNTP and rGTP. Wild-type Taq pol I and A661E demonstrated similar Vmax and Km values for the correct nucleotide dGTP. However, A661E discriminated between incorrect and correct nucleotide less well than wild-type; discrimination was reduced by factors of 9.5-, 5.6- and 15-fold for dATP, dTTP and rGTP, respectively. These data suggest that A661E is efficient polymerases in the presence of the correct deoxynucleotide, dGTP, but it is impaired in ability to discriminate between correct and incorrect deoxyribonucleotides or between ribo- and deoxyribonucleotides. A structural model of Taq pol I is described in which the mutation A661E alters the interactions between the O-helix and the terminal two phosphate groups in the primer strand.

本文言語English
ページ(範囲)197-207
ページ数11
ジャーナルMutation Research - DNA Repair
485
3
DOI
出版ステータスPublished - 04-04-2001
外部発表はい

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Toxicology
  • Genetics

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