Properties of the duplex DNA-dependent ATPase activity of Escherichia coli recA protein and its role in branch migration

S. C. Kowalczykowski, J. Clow, R. A. Krupp

Research output: Contribution to journalArticle

65 Scopus citations

Abstract

We have investigated the double-stranded DNA (dsDNA)-dependent ATPase activity of recA protein. This activity is distinguished from the single-stranded DNA (ssDNA)-dependent ATPase activity by the presence of a pronounced lag time before the onset of steady-state ATP hydrolysis. During the lag phase there is little ATP hydrolysis. The duration of the lag phase, referred to as the lag time, is found to increase with the thermal stability of the dsDNA substrate. Increasing either the MgCl2 or NaCl concentration increases the lag time, whereas increasing the temperature decreases the lag time. The lag time shows little dependence on recA protein concentration but is strongly dependent on ATP concentration. After the lag phase, a steady-state ATP hydrolysis rate is achieved that approaches the rate observed with ssDNA. The steady-state phase of the reaction is proportional to the concentration of recA protein-DNA complex and shows saturation behavior at ~5 ± 1 base pairs per recA protein monomer. These results suggest that the lag phase represents a rate-limiting step in the dsDNA-dependent ATP hydrolysis reaction that requires a structural transition in the dsDNA and that involves a ternary complex of ATP, recA protein, and DNA. We propose that this transition involves the transient denaturation of the dsDNA to form regions of ssDNA. Elsewhere we demonstrate that the dsDNA-dependent ATPase activity is proportional to the rate of recA protein-catalyzed branch migration. We suggest that this activity is responsible for a polar polymerization that drives the branch migration reaction.

Original languageEnglish (US)
Pages (from-to)3127-3131
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume84
Issue number10
StatePublished - 1987
Externally publishedYes

ASJC Scopus subject areas

  • General
  • Genetics

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