The RecA protein from Escherichia coli is a multifunctinal, oligomeric enzyme which is the central catalytic and regulatory component of the processes of recombinational DNA repair and homologous genetic recombination. The goal of our research is to provide a molecular description of the catalytic organization and allosteric mechanism of RecA for both its recombination and coprotease functions. RecA binds cooperatively to single stranded DNA to form helical nucleoprotein filaments which are the active species for both functions. The importance of such a protein structure for the catalysis of recombination has been established with the discovery of RecA-like proteins in all organisms studied to date, from bacteria to humans. Thus, our work with the more tractable bacterial system will provide information relevant to all members of this class of enzymes. We have created large numbers of recA mutants with single or multiple substitutions in targeted domains of the protein structure. Genetic analysis and initial biochemical studies of purified mutant proteins have provided important data regarding both the chemical and steric constraints at specific positions in these domains. Our work will now shift to a more biochemical and structural focus for the analysis of purified mutant RecA proteins in order to correlate the genetic-data with specific molecular defects in RecA function or structure. This work will also provide valuable insights into the molecular mechanisms by which allosterically regulated enzymes transmit information across protein structures.
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