The roles of the Pseudomonas aeruginosa Prc/AlgO protease - Resubmission - 1

项目摘要 细菌病原体铜绿假单胞菌引起危及生命的人类疾病，包括急性 肺炎、囊性纤维化患者的长期肺定植和严重伤口感染。大多数感染 与宿主防御受损有关，这与常见的环境事件一起， 铜绿假单胞菌，使其成为医院获得性感染的主要原因。铜绿假单胞菌病的治疗 通常具有挑战性，部分原因是其对抗生素的内在耐药性以及偶尔爆发的 多重耐药菌株因此，迫切需要表征用于治疗的新靶点。 攻击铜绿假单胞菌细胞被膜含有名为CtpA的C-末端加工蛋白酶（CTP），其 对急性感染至关重要。CtpA的作用与大肠杆菌中唯一的CTP相似 coli、Prc，尽管CtpA和Prc彼此差异很大。铜绿假单胞菌中的CtpA和 E.大肠杆菌，降解重塑细菌细胞壁的酶。这很有趣，因为许多细菌 如E.大肠杆菌只有一个CTP，铜绿假单胞菌有两个，第二个被命名为Prc，因为它非常 与E.大肠杆菌Prc.这就为这一探索性/开发性提案提出了新的问题：如果CtpA 实现了中华人民共和国在E.大肠杆菌，那么中国在宾夕法尼亚州做什么？有人建议中华人民共和国打破 负调节蛋白MucA，触发胞外多糖产生和表型 与囊性纤维化患者预后不良相关。然而，MucA或任何其他铜绿假单胞菌 蛋白质已被证明是Prc底物。根据我们的初步数据，我们提出了一个令人兴奋的 Prc和CtpA对至少一种PG交联水解酶的差异控制的新模型。一起 我们的其他观察表明Prc影响细胞壁，这支持了我们的中心假设， Prc切割参与细胞壁代谢的重要蛋白质，这可能间接影响细胞壁代谢。 对抗σ因子MucA的切割的影响。为了检验这一假设，我们将：（1）确定Prc和 CtpA在降解常见底物中发挥不同的作用;（2）建立Prc如何影响藻酸盐调节 和粘液转化表型。这项工作将影响我们对基本的，保守的 对几乎所有细菌都很重要。它还将提供第一个明确的洞察究竟是什么中国在 PA，以及其影响藻酸盐生物合成、类粘蛋白转化和细胞壁特征的机制 影响人类健康。了解这些机制可能最终有助于开发新的 针对这种广泛的、危险的和非常昂贵的人类病原体的治疗策略。

PROJECT SUMMARY The bacterial pathogen Pseudomonas aeruginosa causes life-threatening human illnesses, including acute pneumonia, long-term lung colonization in cystic fibrosis patients, and severe wound infections. Most infections are associated with compromised host defenses and this, together with the common environmental occurrence of P. aeruginosa, makes it a leading cause of hospital-acquired infections. Treatment of P. aeruginosa disease is often challenging, in part because of its intrinsic resistance to antibiotics as well as occasional outbreaks of multi-drug-resistant strains. Therefore, there is an urgent need to characterize new targets for therapeutic attack. The P. aeruginosa cell envelope contains a C-terminal processing protease (CTP) named CtpA, which is essential for acute infection. CtpA has adopted a similar role to that of the only CTP found in Escherichia coli, Prc, even though CtpA and Prc are quite different from each other. Both CtpA in P. aeruginosa, and Prc in E. coli, degrade enzymes that remodel the bacterial cell wall. This is intriguing because whereas many bacteria like E. coli have only one CTP, P. aeruginosa has two, and the second one is named Prc because it is very similar to E. coli Prc. This raises the new question for this exploratory/developmental proposal: if CtpA achieves the role played by Prc in E. coli, then what does Prc do in Pa? Prc has been proposed to cleave the negative regulatory protein MucA, triggering extracellular polysaccharide production and a phenotype associated with a poor prognosis in cystic fibrosis patients. However, neither MucA or any other P. aeruginosa protein has been demonstrated to be a Prc substrate. From our preliminary data, we are proposing an exciting new model for the differential control of at least one PG cross-link hydrolase by both Prc and CtpA. Together with our other observations suggesting that Prc affects the cell wall, this supports our central hypothesis, which is that Prc cleaves important proteins involved in cell wall metabolism, and that this might have an indirect impact on the cleavage of the anti-sigma factor MucA. To test this hypothesis, we will: (1) Determine if Prc and CtpA play different roles in degrading common substrates and (2) Establish how Prc affects alginate regulation and the mucoid conversion phenotype. This work will impact our understanding of fundamental, conserved processes important to almost all bacteria. It will also provide the first clear insight into exactly what Prc does in Pa, and the mechanism(s) by which it affects alginate biosynthesis, mucoid conversion, and cell wall features that impact human health. Understanding these mechanisms might ultimately help the development of new therapeutic strategies against this widespread, dangerous and very costly human pathogen.