Differential rates of NTP hydrolysis by the mutant [S69G]RecA protein. Evidence for a coupling of NTP turnover to DNA strand exchange

Sunil Nayak, Floyd R. Bryant

Research output: Contribution to journalArticlepeer-review

Abstract

The x-ray crystal structure of the Escherichia coli RecA protein indicates that the phosphate groups of the nucleotide cofactor are bound by a loop whose amino acid sequence (66GPESSGKT73) corresponds to a consensus phosphate binding loop sequence (GXXXXGK[T/S]) found in many NTP-binding proteins. As part of an investigation of the role of the P-loop in ATP hydrolysis, we prepared a mutant RecA protein in which serine 69 was replaced by a glycine residue. We have found that the [S69G]RecA mutation has a differential effect on the hydrolysis of various nucleoside triphosphates. The [S69G]RecA protein catalyzes the single-stranded DNA-dependent hydrolysis of rATP, ddATP, and dATP with turnover numbers of 10, 20, and 36 min-1, respectively. The wild type RecA protein, in contrast, hydrolyzes each of these nucleoside triphosphates with similar turnover numbers of 20-24 min- 1. Significantly, the [S69G]RecA protein promotes strand exchange with all three nucleoside triphosphates, and the rate of strand exchange is directly proportional to the rate of hydrolysis of each of the nucleotide cofactors. These findings with the [S69G]RecA protein provide support for the existence of a mechanistic coupling between NTP hydrolysis and DNA strand exchange.

Original languageEnglish (US)
Pages (from-to)25979-25982
Number of pages4
JournalJournal of Biological Chemistry
Volume274
Issue number37
DOIs
StatePublished - Sep 10 1999

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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