ATP Hydrolysis promotes duplex DNA release by the RecA presynaptic complex

Abstract

Homologous recombination is an important DNA repair pathway that plays important 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 singlestranded DNA, called presynaptic complexes, which are responsible for locating and pairing with a homologous duplex DNA. Extensive studies have been done to reveal the roles of ATP hydrolysis of RecA in the reactions, but its potential influence remains poorly understood. We investigate how ATP hydrolysis influences the interaction between RecA presynaptic complex and homologous dsDNA. We demonstrate that over short time regimes, RecA presynaptic complexes sample heterologous dsDNA similarly regardless of 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 about 4-times higher than with ATPγS. These experiments suggest that ATP hydrolysis 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.