Bruns O, Bruns W, Pulverer G
Institut für Pharmakologie, Universität zu Köln, Germany.
Zentralbl Bakteriol. 1997 Feb;285(3):413-30. doi: 10.1016/s0934-8840(97)80008-4.
Nearly all clinical isolates of methicillin resistant Staphylococcus aureus (MRSA) produce beta-lactamase as well as an additional low-affinity penicillin-binding protein called PBP2a or PBP2', the main factor for mediating methicillin resistance. Polidocanol (PDO), a dodecyl polyethyleneoxide ether, resensitizes clinical isolates of MRSA to methicillin; in addition, their resistance to benzylpenicillin (BP) is reduced. The action of PDO is based on the inhibition of the induced syntheses of PBP2a and beta-lactamase. Induction in our study was performed with 2-(2'-carboxyphenyl)benzoyl-6-aminopenicillanic acid (CBAP). Inducible PBP2a production in MRSA strains is under the control of the same regulatory system which is responsible for the induction of beta-lactamase synthesis. BlaR1, a membrane-spanning protein with a penicillin sensor and a signal transducer domain represents the starting point of this induction cascade. Based on its amphiphilic properties, it is likely that the action of PDO is located in the bacterial membrane. Therefore we investigated the possibility that BlaR1 might be the main target for PDO action. We were able to detect the BlaR1 sensor domain in resistant staphylococcal cells even in the noninduced state by fluorography. In a competition assay, CBAP was bound specifically, with a high affinity to the penicillin sensor. Moreover, the binding of CBAP was very stable. As concerns PDO, no significant interaction with the penicillin binding site of BlaR1 was detectable. This is why the BlaR1 transducer domain is thought to be the actual target area of PDO. In this case, PDO would interfere with the transmission of the signal, generated by the receptor binding of CBAP, through the membrane via BlaR1 into the staphylococcal cell. This assumption could be confirmed by the analysis of the concentration-effect relationship, whereafter PDO does not work as a competitive, but as a noncompetitive antagonist of CBAP. Our results demonstrate that BlaR1 could be an attractive new target for the development of new drugs to overcome methicillin resistance.
几乎所有耐甲氧西林金黄色葡萄球菌（MRSA）临床分离株都能产生β-内酰胺酶以及另一种名为PBP2a或PBP2'的低亲和力青霉素结合蛋白，这是介导甲氧西林耐药性的主要因素。聚多卡醇（PDO），一种十二烷基聚环氧乙烷醚，可使MRSA临床分离株对甲氧西林重新敏感；此外，它们对苄青霉素（BP）的耐药性也会降低。PDO的作用基于对PBP2a和β-内酰胺酶诱导合成的抑制。在我们的研究中，诱导是用2-（2'-羧基苯基）苯甲酰基-6-氨基青霉烷酸（CBAP）进行的。MRSA菌株中可诱导的PBP2a产生受负责β-内酰胺酶合成诱导的同一调节系统控制。BlaR1是一种具有青霉素传感器和信号转导结构域的跨膜蛋白，是这种诱导级联反应的起点。基于其两亲性，PDO的作用可能位于细菌膜中。因此我们研究了BlaR1可能是PDO作用主要靶点的可能性。通过荧光自显影，我们甚至在未诱导状态下也能在耐药葡萄球菌细胞中检测到BlaR1传感器结构域。在竞争试验中,CBAP能以高亲和力特异性结合到青霉素传感器上。此外，CBAP的结合非常稳定。至于PDO，未检测到其与BlaR1的青霉素结合位点有明显相互作用。这就是为什么BlaR1转导结构域被认为是PDO的实际作用靶点区域。在这种情况下，PDO会干扰由CBAP受体结合产生的信号通过BlaR1经膜传入葡萄球菌细胞的传递过程。这一假设可以通过浓度效应关系分析得到证实，结果显示PDO不是作为CBAP的竞争性拮抗剂，而是作为非竞争性拮抗剂起作用。我们的结果表明，BlaR1可能是开发克服甲氧西林耐药性新药的一个有吸引力的新靶点。
