Inducible Antibiotic Resistance in Methicillin-Resistant Staphylococcus Aureus

描述(申请人提供)：耐甲氧西林金黄色葡萄球菌(MRSA)已经获得了对内酰胺类抗生素的可诱导耐药机制，基本上涵盖了抗生素类别的所有成员。这种耐药性是由一组编码抗生素传感器/信号转导蛋白、基因抑制子和两个耐药决定因素的基因授予的，A类内酰胺酶和一种特殊的青霉素结合蛋白(PBP)被称为PBP2a。我们已经证明，抗生素传感器/信号转导蛋白BlaR1在其膜表面区经历了β-内酰胺类抗生素的共价修饰，这一过程通过我们称为“赖氨酸N-脱羧基开关”的独特过程激活蛋白质，以进行跨膜的信号转导。在这一事件之后，BlaR1的细胞质结构域经历了磷酸化，所有这些都在与诱导抗性相关的时间框架内。阐明这种BlaR1磷酸化对抗生素耐药事件的重要性是在特定目标1下的研究主题。PBP2a在MRSA中执行细胞壁的交联性，这是其生存所必需的功能。由于PBP2a具有封闭的活性部位，不能被内酰胺类抗生素很好地抑制，因此它在耐药中发挥作用。我们已经阐明了该蛋白上的一个变构位点，它被触发以促进该蛋白的生理作用的活性部位的打开。变构位点是PBP2a的阿切莱斯后跟，因为它触发活性部位的开放会使蛋白质(和MRSA)容易受到内酰胺类抗生素的攻击，而这些抗生素在MRSA感染的治疗中已经过时。在特定的目标2中，我们建议研究这种蛋白如何发挥其生理作用，以及它的过程如何在设计治疗MRSA感染的新策略时被颠覆。此外，我们建议研究由变构位点改变引起的抗生素耐药机制。

DESCRIPTION (provided by applicant): Methicillin-resistant Staphylococcus aureus (MRSA) has acquired an inducible resistance mechanism to ¿- lactam antibiotics that encompasses essentially all members of the antibiotic class. This resistance is conferred by a set of genes that encode an antibiotic sensor/signal transducer protein, gene repressor and two resistant determinants, a class A ¿-lactamases and a special penicillin-binding protein (PBP) referred to as PBP2a. We have documented that the antibiotic sensor/signal transducer protein BlaR1 experiences covalent modification by ¿-lactam antibiotics in its membrane-surface domain, which through a unique process that we have termed "lysine N-decarboxylation switch" activates the protein for signal transduction across the membrane. Subsequent to this event, the cytoplasmic domain of BlaR1 experiences phosphorylation, all within the time frame relevant to induction of resistance. The elucidation of the importance of this BlaR1 phosphorylation to the antibiotic resistance events is the subject of study under Specific Aim 1. PBP2a performs cross-linking of the cell wall in MRSA, a function that is indispensible to its survival. PBP2a is not inhibited well by ¿-lactam antibiotics as it has a closed active site, hence its function in resistance. We have elucidated an allosteric site on this protein that is triggered to facilitate opening of the active site for the physiological role of the protein. The allosteric site is an Achiles' Heel of PBP2a, since its triggering for the opening of the active site would leave the protein (and MRSA) vulnerable to ¿-lactam antibiotics that have met their obsolescence in treatment of infections by MRSA. In Specific Aim 2 we propose to investigate how this protein performs its physiological role and how its processes can be subverted in devising new strategies in treatment of MRSA infections. Furthermore, we propose to study antibiotic resistance mechanisms that arise by alterations in the allosteric site.