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J Biol Chem. 2016 Oct 14;291(42):22218-22230. Epub 2016 Sep 1.

ATP hydrolysis Promotes Duplex DNA Release by the RecA Presynaptic Complex.

Author information

1
From the Department of Biochemistry & Molecular Biophysics, Columbia University, New York, New York 10032 and.
2
the Center of Quantitative Biology & Center of Life Sciences, Academy for Advanced Interdisciplinary Studies, Peking University, Beijing, 100871 China.
3
From the Department of Biochemistry & Molecular Biophysics, Columbia University, New York, New York 10032 and ecg2108@cumc.columbia.edu.

Abstract

Homologous recombination is an important DNA repair pathway that plays key roles in maintaining genome stability. Escherichia coli RecA is an ATP-dependent DNA-binding protein that catalyzes the DNA strand exchange reactions in homologous recombination. RecA assembles into long helical filaments on single-stranded DNA, and these presynaptic complexes are responsible for locating and pairing with a homologous duplex DNA. Recent single molecule studies have provided new insights into RecA behavior, but the potential influence of ATP in the reactions remains poorly understood. Here we examine how ATP influences the ability of the RecA presynaptic complex to interact with homologous dsDNA. We demonstrate that over short time regimes, RecA presynaptic complexes sample heterologous dsDNA similarly in the presence of either ATP or ATPγS, suggesting that initial interactions do not depend on ATP hydrolysis. In addition, RecA stabilizes pairing intermediates in three-base steps, and stepping energetics is seemingly unaltered in the presence of ATP. However, the overall dissociation rate of these paired intermediates with ATP is ∼4-fold higher than with ATPγS. These experiments suggest that ATP plays an unanticipated role in promoting the turnover of captured duplex DNA intermediates as RecA attempts to align homologous sequences during the early stages of recombination.

KEYWORDS:

ATP; DNA recombination; DNA repair; homologous recombination; single-molecule biophysics

PMID:
27587394
PMCID:
PMC5064001
DOI:
10.1074/jbc.M116.740563
[Indexed for MEDLINE]
Free PMC Article

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