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）已经获得了对内酰胺类抗生素的诱导耐药机制，该机制基本上涵盖了抗生素类的所有成员。这种耐药性是由一组基因赋予的，这些基因编码一种抗生素传感器/信号传感器蛋白、基因抑制因子和两种耐药决定因子，即a类内酰胺酶和一种称为PBP2a的特殊青霉素结合蛋白（PBP）。我们已经记录了抗生素传感器/信号传感器蛋白BlaR1在其膜表面区域经历了内酰胺抗生素的共价修饰，这是通过一个独特的过程，我们称之为“赖氨酸n -脱羧开关”，激活蛋白质进行跨膜信号转导。在此事件之后，BlaR1的细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框架内。阐明BlaR1磷酸化对抗生素耐药事件的重要性是Specific Aim 1研究的主题。PBP2a在MRSA中执行细胞壁交联，这是其生存所必需的功能。由于PBP2a具有一个封闭的活性位点，因此它在耐药中起作用，因此不被内酰胺类抗生素很好地抑制。我们已经阐明了该蛋白上的一个变构位点，该位点被触发以促进该蛋白生理作用的活性位点的打开。变构位点是PBP2a的致命弱点，因为它触发活性位点的打开会使蛋白质（和MRSA）容易受到内酰胺类抗生素的攻击，而内酰胺类抗生素在治疗MRSA感染方面已经过时了。在Specific Aim 2中，我们建议研究这种蛋白如何发挥其生理作用，以及在设计治疗MRSA感染的新策略时如何颠覆其过程。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.