PROTEASE SECRETION IN PSEUDOMONAS AERUGINOSA

简介(摘自申请者摘要)：铜绿假单胞菌 一种主要的机会性病原体，本研究将阐明其致病机理 机械装置。铜绿假单胞菌分泌大量有毒物质的能力 降解酶在其发病机制中起主要作用。其中包括 胞外酶是LasB酶(弹性酶)和LasA酶(溶葡萄球菌 蛋白酶)协同作用，降解弹性蛋白和许多其他 临床上相关的蛋白质。LasA和LasB酶最初都是形成的 作为具有三个结构域的前酶原：信号序列、前肽和 成熟层序。前肽结构域起分子内伴侣的作用， 是酶的活性和细胞外定位所必需的。 这项建议的目的是了解LasA和LasB分子 结构，其伴侣辅助折叠的机制，途径 铜绿假单胞菌分泌物与临床相关性。这项建议结合了 分子、生化和生物物理方法比较加工和 分泌这两种重要的蛋白酶。调查人员将分析 LasA和LasB外源蛋白的遗传和结晶学结构 方法：研究方法。他们将确定前肽(伴侣)介导的机制 LasA和LasB蛋白的折叠和分泌。他们将决定 ProLasA和ProLasB与II型分泌物相互作用的机制 释放到环境中的设备。他们还将确定临床上的 LASA和LasB在发病机制中的相关性，并检验LASA作为一种 特殊的抗葡萄球菌治疗。这项研究将会得出 在新的信息中对重要的发病机制进行了重要的 机会主义有机体。它还提供了一个模型系统，用于理解 革兰氏阴性菌蛋白质输出的主要途径。

DESCRIPTION (Adapted from the Applicant's Abstract): Pseudomonas aeruginosa is a major opportunistic pathogen, and this study will elucidate its pathogenic mechanisms. The ability of P. aeruginosa to secrete numerous toxic and degradative enzymes plays a major role in its pathogenesis. Among these exoenzymes are LasB protease (elastase) and LasA protease (staphylolytic protease) which act synergistically to degrade elastin and many other clinically relevant proteins. Both LasA and LasB proteases are initially formed as pre-proenzymes with three domains: a signal sequence, a propeptide and the mature sequence. The propeptide domain acts as an intramolecular chaperone that is required for both the enzyme's activity and its extracellular localization. The objective of this proposal is to understand LasA and LasB molecular structures, the mechanisms of their chaperone-assisted folding, pathways of secretion in P. aeruginosa, and clinical relevance. This proposal combines molecular, biochemical and biophysical approaches to compare the processing and secretion of these two important proteases. The investigators will analyze the structures of LasA and LasB exoproteins by genetic and crystallographic methods. They will determine the mechanisms of propeptide (chaperone)-mediated folding and secretion of LasA and LasB proteins. They will determine the mechanism by which proLasA and proLasB interact with the type II secretion apparatus for release to the environment. They will also determine the clinical relevance of LasA and LasB in pathogenesis, and test the efficacy of LasA as a specific anti-staphylococcal therapeutic treatment. This research will result in new information on a significant mechanism of pathogenesis in an important opportunistic organism. It also provides a model system for understanding a major pathway of protein export in gram-negative bacteria.