Conserved complement binding to bacterial porins

描述（由申请人提供）：本申请旨在定义补体C3与细菌表面结合相互作用的分子机制。具体来说，我们将评估如何铜绿假单胞菌C3结合到外膜孔蛋白（OprF）促进吞噬细胞的相互作用，以及如何利用这可能会提高调理吞噬和补体介导的杀伤。我们还发现铜绿假单胞菌OprF的C-末端区域与几种其他革兰氏阴性细菌的OmpA结构域具有高度的结构同一性，已知其中一种细菌结合C3。因此，这项研究不仅有助于了解铜绿假单胞菌的发病机制，而且还为其他革兰氏阴性菌中补体相互作用的生物学提供了见解。这将反过来导致新的和更有效的策略，用于治疗或预防铜绿假单胞菌和其他革兰氏阴性菌感染。该提案将使用最先进的分子，生物化学和遗传学方法来探测补体和革兰氏阴性孔蛋白的相互作用。迄今为止，对补体介导的铜绿假单胞菌杀伤的抗性的机制基础尚未完全理解。我们最近发现OprF是人补体C3识别的一种配体。目的1将评价铜绿假单胞菌临床分离株中OprF-C3结合和Psl掩蔽的保守性。目的2的目标是确定OprF水平是否可以被调节以改善血清介导的杀伤。目标3将侧重于映射C3相互作用所需的OprF区域。最后，在目标4中，我们将评估C3相互作用的守恒性 在其它革兰氏阴性病原体中具有表面暴露的孔蛋白结构域。了解C3与细菌表面结合的分子机制为预防和/或治疗由这些革兰氏阴性病原体引起的疾病提供了潜在的手段。

DESCRIPTION (provided by applicant): This application seeks to define the molecular mechanism of complement C3 binding interactions with bacterial surfaces. Specifically we will evaluate how Pseudomonas aeruginosa C3 binding to an outer membrane porin (OprF) promotes phagocyte interactions and how this may be exploited to enhance opsonophagocytosis and complement-mediated killing. We also discovered that the C-terminal region of P. aeruginosa OprF has high structural identity with the OmpA domain of several other Gram- negative bacteria, one of which is known to bind C3. Therefore, this study will not only contribute to an understanding of P. aeruginosa pathogenesis but also provide insights into the biology of complement interactions in other Gram-negative bacteria. This will, in turn, lead to new and more effective strategies for the treatment or prevention of P. aeruginosa and perhaps other Gram-negative infections. This proposal will use state-of-the art molecular, biochemical, and genetic approaches to probe interactions of complement and Gram-negative porins. To date, the mechanistic basis for resistance to complement-mediated killing of P. aeruginosa is not completely understood. We recently discovered that OprF is one ligand recognized by human complement C3. Aim 1 will evaluate conservation of OprF-C3 binding and Psl masking among P. aeruginosa clinical isolates. The goal of Aim 2 is to determine if OprF levels can be modulated to improve serum-mediated killing. Aim 3 will focus on mapping regions of OprF that are required for C3 interactions. Finally, in Aim 4 we will evaluate conservation of C3 interaction with surface exposed porin domains in other Gram-negative pathogens. Understanding the molecular mechanism of C3 binding to bacterial surfaces offers a potential means for preventing and/or treating diseases caused by these Gram-negative pathogens.