Structure/function of RecA protein from P aeruginosa

大肠杆菌RecA蛋白在体外促进DNA链交换反应 这为以下中心步骤提供了一个方便的分子模型 重组DNA修复和同源基因重组。长距离的 ROL GM32335(COX)的研究目标是详细了解 RecA介导的DNA链交换。重组DNA的假说 修复是RecA蛋白在体内的主要功能提供智力 框架。 GM32335-17(最近资助)的具体目标之一是执行 细菌RecA蛋白和RecA同源物的比较分析。 铜绿假单胞菌是一种重要的人类病原体，是一种 待研究的RecA蛋白。在本提案中，一个协作 这将大大扩展我们提出的对假单胞菌的分析 铜绿假单胞菌RecA蛋白，并改善机制和前景 结构洞察力。与弗拉迪斯拉瓦·兰佐夫博士的实验室一起 俄罗斯圣彼得堡，我们将首先探索生物化学 铜绿假单胞菌RecA蛋白的表达，并建立其与 电子线圈RecA。使用广泛的活性嵌合蛋白 由两种RecA蛋白的融合构建而成，我们将试图精确定位 造成这种差异的蛋白质区域。这项工作应该会有帮助 测试当前RecA介导的DNA链交换模型的关键特征，以及 也可能有助于识别具有增强的DNA结合和链的RecA变体 交换功能。这样的蛋白质最终可能会被证明是有用的 基因治疗方案中的RecA和产生RecA-DNA复合体的晶体 用于结构分析。

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.