Cell-Wall Recycling and Nexus to Antibiotic Resistance

项目摘要/摘要 细胞壁回收是细菌的一个基本过程，通过它可以重塑细菌的细胞壁。 在正常生长的过程中，以及对抗生素的反应，抗生素对细胞壁造成破坏 行动机制。肠杆菌科和铜绿假单胞菌(本次拨款申请的对象) β-内酰胺类抗生素对其细胞壁造成的感官损伤。感应事件与细胞壁相连。 循环，这会导致以细胞壁为基础的天然产品的形成，即所谓的神经肽。一定的 肌肽内化到细胞质中，在那里它们诱导细菌对抗生素的反应。 (抗生素耐药机制)。这一过程导致了许多β-内酰胺类抗生素的过时 对抗革兰氏阴性细菌。我的实验室在过去的几年里一直在研究这个系统，我披露的是 在这个Mira应用程序中，是实验室将在不久的将来绘制的路径。我建议研究一下 含有裂解转糖基酶(LTS)的周质复合体，它翻转细胞壁以达到 回收利用或对抗生素造成的损害作出反应。我的实验室记录了P中有11个已知的LT。 铜绿假单胞菌，我们已经描述了它与细胞壁的个别反应。这些酶是 建议参与周质内其他蛋白质的复合体，其身份尚不清楚 这代表了我们对细胞壁过程的认识上的一个重大空白。而LTS的所有功能并不都是 据了解，一种是细菌暴露于β-内酰胺类抗生素后细胞壁的修复。多肽是 细胞壁循环的降解产物，为此目的内化到细胞质中。作为一种 在循环事件的分支，某些神经肽激活AMPR转录调节因子的表达 AmpC酶β-内酰胺酶，β-内酰胺类抗生素的耐药决定因素。我们将研究它们之间的相互作用 AMPR蛋白在阐明该系统中的关键多肽。此外，还增加了四个 已在铜绿假单胞菌中确定的与肌肽关键事件相关的细胞质酶 将对它们的化学反应、催化循环的细节以及它们的回收进行调查 结构。我预计，成功完成这项拟议的科学不仅将导致 阐明这些调节细胞壁的重要事件，也将为他们的 阻断作为一种规避革兰氏阴性β-内酰胺类耐药复杂机制的手段 细菌已经进化了。人们对这些复杂的事件知之甚少，它们将在我的 实验室正在进行拟议的研究。

Project Summary/Abstract Cell-wall recycling is a fundamental process in bacteria, whereby it allows for remodeling of the cell wall in the course of the normal growth and in response to antibiotics that inflict damage to the cell wall for their mechanisms of action. Enterobacteriaceae and Pseudomonas aeruginosa (subject of this grant application) sense damage inflicted to their cell wall by β-lactam antibiotics. The sensing event is linked to cell-wall recycling, which leads to the formation of cell-wall-based natural products known as muropeptides. Certain muropeptides are internalized to the cytoplasm, where they induce the bacterial response to the antibiotic (antibiotic-resistance mechanisms). This process has led to obsolescence of many of the β-lactam antibiotics against Gram-negative bacteria. My lab has studied this system for the past several years and what I disclose in this MIRA application is the path that the lab will chart in the immediate future. I propose to study the periplasmic complexes involving lytic transglycosylases (LTs), which turn over the cell wall for the purpose of recycling or in response to damage by antibiotics. My lab has documented that there are 11 known LTs in P. aeruginosa, whose individual reactions with the cell wall have been described by us. These enzymes are proposed to be involved in complexes with other proteins within the periplasm, whose identities are not known and represent a major gap in our knowledge of cell-wall processes. Whereas all the functions of LTs are not understood, one is repair of cell wall upon exposure of bacteria to β-lactam antibiotics. Muropeptides are the degradation products of cell-wall recycling, which are internalized to the cytoplasm for this purpose. As an offshoot of the recycling events, certain muropetides activate the AmpR transcriptional regulator in expression of the AmpC β-lactamase, the resistance determinant for β-lactam antibiotics. We will study the interactions of the key mutropeptides with the AmpR protein in elucidating the system. Furthermore, four additional cytoplasmic enzymes that have been identified in P. aeruginosa for the key events of the muropeptide recycling will be investigated for their chemical reactions, the details of the catalytic cycles and for their structures. I anticipate that the successful completion of this proposed science will not only lead to the elucidation of these important events regulating the cell wall, but also will identify opportunities for their interruption as a means to circumventing the elaborate mechanism of β-lactam resistance that Gram-negative bacteria have evolved. These complex events are poorly understood, and they will be studied in detail in my lab in the course of the proposed reseach.