Inducible Antibiotic Resistance in Methicillin-Resistant Staphylococcus Aureus

描述（由申请人提供）：耐甲氧西林金黄色葡萄球菌（MRSA）已获得对ε-内酰胺类抗生素的诱导性耐药机制，基本上包括抗生素类的所有成员。这种耐药性是由一组编码抗生素传感器/信号转导蛋白、基因阻遏物和两种耐药决定簇（A类内酰胺酶和称为PBP 2a的特殊青霉素结合蛋白（PBP））的基因赋予的。我们已经记录了抗生素传感器/信号转导蛋白BlaR 1在其膜表面结构域中经历了<$-内酰胺抗生素的共价修饰，通过我们称为“赖氨酸N-脱羧开关”的独特过程激活蛋白质进行跨膜信号转导。在此事件之后，BlaR 1的胞质结构域经历磷酸化，所有这些都在与诱导抗性相关的时间范围内。阐明这种BlaR 1磷酸化对抗生素耐药性事件的重要性是特定目标1下的研究主题。PBP 2a在MRSA中执行细胞壁的交联，这是其生存所不可或缺的功能。由于PBP 2a具有封闭的活性位点，因此其在耐药性中起作用，因此不能很好地被<$-内酰胺类抗生素抑制。我们已经阐明了这种蛋白质上的变构位点，其被触发以促进蛋白质的生理作用的活性位点的开放。变构位点是PBP 2a的一个“脚后跟”，因为它触发活性位点的开放将使蛋白质（和MRSA）易受内酰胺类抗生素的攻击，这些抗生素在治疗MRSA感染时已经过时。在具体目标2中，我们建议研究这种蛋白质如何发挥其生理作用，以及如何在设计治疗MRSA感染的新策略时颠覆其过程。此外，我们建议研究由变构位点的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.