BINDING OF PHOSPHOLIPASE A2 TO BILAYERS

磷脂酶A2对磷脂双层和生物膜的作用 取决于他们的组织。这项研究的实验目的是 表征磷脂酶A2对阴离子囊泡的作用 为了了解阴离子表面的作用，两性离子脂 电荷的作用，底物结合到活性部位的作用 界面识别部位，以及钙结合部位的作用。从… 我们也希望通过这些实验来加深对 不同结合部位与动力学之间的相互关系 参数。因此，例如，酶在小泡内的滑行导致了 更高的初始水解率，而更快的小泡间交换 结合酶的浓度会降低表观水解率。 我们还计划通过以下方法来表征结合酶的微环境 探测双层中的酶的状态，并通过监测相 在存在和不存在的情况下衬底双层的状态 酵素。双层的相态和局部扰动将是 特点是监测跨双层运动，标志物释放，侧向 相分离，囊泡融合。这些实验是经过精心设计的 以衬底界面中的缺陷调节 结合和酶的催化周转。 这些实验预计将提供关于结合和 底物结构和相态的催化专一性 它们的双层，关于结合酶的跨囊泡交换的速率， 关于相态对动力学参数和结合参数的影响。 这一信息是理解界面机制的核心。 磷脂酶A2的激活。激活和特异度 磷脂酶A2被认为是第一步也是速度限制步骤 从磷脂酶A2开始前列腺素的体内生物合成 从膜中释放或调节花生四烯酸的释放 磷脂。所有这些研究都将在磷脂酶A2上进行 从猪的胰腺中提取，并可能在以后从其他 消息来源。

Action of phospholipase A2 on phospholipid bilayers and biomembranes depends upon their organization. The experimental aim of this study is to characterize the action of phospholipase A2 on vesicles of anionic and zwitterionic lipids in order to understand the role of anionic surface charge, the role of substrate binding to the active site, the role of interface recognition site, and the role of calcium binding sites. From these experiments we also hope to develop an understanding of the interrelationships between the various binding sites and kinetic parameters. Thus, for example, intravesicle scooting of the enzyme leads to higher initial rate of hydrolysis, whereas faster intervesicle exchange of the bound enzyme would lower apparent rate of hydrolysis. We also plan to characterize the microenvironment of the bound enzyme by probing the state of the enzyme in the bilayer, and by monitoring the phase state of the substrate bilayer in the presence and in the absence of the enzyme. The phase state and local purturbation of the bilayer will be characterized by monitoring transbilayer movement, marker release, lateral phase separation, and fusion of vesicles. These experiments are designed with the paradigm that defects in the substrate interface regulate the binding and the catalytic turnover of the enzyme. Such experiments are expected to provide information about binding and catalytic specificity for the substrate structure and the phase state of their bilayer, about rates of transvesicle exchange of the bound enzyme, about the effect of the phase state on the kinetic and binding parameters. This information is central to understanding the mechanism of interfacial activation of phospholipase A2. Activation and specificity of phospholipase A2 is thought to be the first and the rate limiting step in the biosynthesis of prostaglandins in vivo, since the phospholipase A2 releases or regulates the release of arachidonic acid from membrane phospholipids. All these studies will be conducted on phospholipase A2 from pig pancreas, and could be later extended to the enzyme from other sources.