Catalytic definition of insecticide-metabolizing P450s

描述(由申请人提供)：昆虫持续暴露在一系列天然有毒植物化学物质和合成有毒杀虫剂中，这些化学物质用于管理昆虫，传播人类和动物疾病并破坏农作物。在越来越多的案例中，P450介导的解毒机制的进化使昆虫能够代谢这些化合物，并使它们能够在高浓度植物毒素和杀虫剂存在的情况下存活。自20世纪40年代S开始使用合成杀虫剂以来，昆虫对杀虫剂的抗药性已经增加到存在于500多个物种中，其中许多物种对人类健康有重大影响，例如传播疟疾寄生虫的按蚊(估计每年导致多达300万人死亡)，以及传播黄热病和西尼罗河疾病的伊蚊和库蚊。我们对这些抗性的分子基础的理解以及我们抵消这些抗性的能力取决于昆虫P450的综合分子模型的开发，该模型将理论建模与蛋白质表达和定向突变相结合，旨在对这些模型进行生化测试。所描述的项目旨在将我们的P450建模和蛋白质表达工作扩展到描述果蝇和按蚊中杀虫剂代谢P450的催化部位，果蝇是发展杀虫剂抗药性的模式生物，而按蚊是许多人类病原体的重要媒介。在分子模拟水平上，对这些结构的比较可以提供以下信息：能够处理杀虫剂的催化位点的相似性(如果有的话)、排除某些催化位点与杀虫剂结合和代谢的差异以及对每种蛋白质的潜在底物和抑制剂的预测。在蛋白质表达和高通量筛选的水平上，描绘能够与每个催化位点结合的化合物的范围可以定义潜在地能够干扰杀虫剂新陈代谢的化合物，从而能够干扰携带病原体的昆虫对当前杀虫剂的抗性生长。这项分析中使用的分子、生化和理论方法的跨学科混合远远超过了大多数抗药性领域的研究人员，为我们提供了在蚊子物种中识别P450抑制剂的巨大潜力，这些蚊子物种作为疾病媒介对健康构成重大风险。

DESCRIPTION (provided by the applicant): Insects are continually exposed to an array of natural toxic plant chemicals and synthetic toxic insecticides that are used in the management of insects vectoring human and animal diseases and damaging crops. In a growing number of cases, the evolution of P450-mediated detoxification mechanisms have allowed insects to metabolize these compounds and enabled them to survive in the presence of high concentrations of plant toxins and insecticides. From the inception of synthetic insecticide usage in the 1940's, the acquisition of insecticide resistances in insects has increased to the point that they exist in over 500 species with many in species that significantly impact human health, such as the Anopheles mosquitoes that vector malarial parasites (estimated to kill as many as 3 million people per year) and the Aedes and Culex mosquitoes that vector yellow fever and West Nile disease. Our understanding of the molecular basis for these resistances and our ability to counteract these resistances depends on the development of comprehensive molecular models for insect P450s that couple theoretical modeling with protein expression and directed mutagenesis aimed at biochemically testing these models. The projects described are aimed at extending our P450 modeling and protein expression efforts to characterization of the catalytic sites in insecticide-metabolizing P450s in Drosophila, a model organism for the development of insecticide resistance, and in Anopheles, a significant vector for many human pathogens. At the level of molecular modeling, comparisons of these structures can provide information on the similarities (if any) of catalytic sites capable of handling insecticides, the differences that preclude some catalytic sites from binding and metabolizing insecticides and predictions on potential substrates and inhibitors for each protein. At the level of protein expression and high-throughput screening, profiling the range of compounds capable of binding to each catalytic site can define compounds potentially capable of interfering with insecticide metabolisms and, therefore, growth of pathogen-bearing insects resistant to current insecticides. The interdisciplinary mixture of molecular, biochemical and theoretical approaches used in this analysis are well beyond those available to most researchers in the field of insecticide resistance and provide us with significant potential for identifying inhibitors for P450s in mosquito species posing significant health risks as disease vectors.