The recA protein of E. coli promotes a DNA strand exchange reaction in vitro that provides a convenient molecular model for the central steps in homologous recombination and recombinational repair. The primary goal of this proposal is to understand the mechanism of this reaction. Major aspects of the reaction to be studied include: (a) the structure of a putative 3-stranded DNA pairing intermediate, (b) the molecular role of ATP hydrolysis, and (c) the molecular function of the acidic carboxyl-terminus of the recA protein. As part of the effort to examine the role of ATP hydrolysis, a yeast protein that promotes DNA strand exchange without ATP will also be examined. The project should offer insights into a novel DNA structure, cellular energy transductions, and broader questions about the repair function of homologous recombination. The health-relatedness aspects of the work concern the implications for recombinational DNA repair processes in the cell. Recombinational repair is the cellular first line of defense against several classes of potentially mutagenic (and thereby carcinogenic) DNA lesions. These include double-strand breaks, double-strand crosslinks, and lesions left in a single-strand gap after replication bypass. The recombinational repair function of recA and other proteins involved in homologous recombination has received inadequate attention. The project may also offer insights into recombination mechanisms that could facilitate the development of safer and more efficient methods for gene therapy.
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