Structure/function of RecA protein from P aeruginosa

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The RecA protein of E. coli promotes a DNA strand exchange reaction in vitro that provides a convenient molecular model for the central steps of recombinational DNA repair and homologous genetic recombination. The long-range goal of the research in ROl GM32335 (Cox) is a detailed understanding of RecA-mediated DNA strand exchange. The hypothesis that recombinational DNA repair is the primary function of RecA protein in vivo provides an intellectual framework. One of the specific aims of GM32335-17 (recently funded) is to carry out a comparative analysis of bacterial RecA proteins and RecA homologues. Pseudomonas aeruginosa, an important human pathogen, is the source of one of the RecA proteins to be investigated. In the present proposal, a collaboration is proposed that should greatly extend our proposed analysis of the Pseudomonas aeruginosa RecA protein, and improve the prospects for mechanistic and structural insights. Together with the laboratory of Dr. Vladislava Lanzov in St. Petersburg, Russia, we will first explore the biochemistry of the P.aeruginosa RecA protein, and establish enzymatic differences between it and the E. coil RecA. Using an extensive set of active chimeric proteins constructed from fusions of the two RecA proteins, we will try to pinpoint regions of the protein responsible for the differences. The work should help test key features of current models for RecA-mediated DNA strand exchange, and may also help identify RecA variants with enhanced DNA binding and strand exchange functions. Such proteins may eventually prove useful in efforts to use RecA in gene therapy protocols and to generate crystals of RecA-DNA complexes for structural analysis.