ANESTHETIC ALTERATION OF MEMBRANE-PROTEIN INTERACTIONS

我们的假设是膜结构的波动起到了 在调节生物功能方面起着重要作用。假设 麻醉作用的部位是脑组织的脂质基质 生物膜，我们提出了初始事件在 麻醉是这种波动的变化。为了测试这一点 假设我们计划使用凝胶-液晶转变 作为这种波动的模型的单双层囊泡和 探讨不同麻醉剂对融化的影响 温度和平均簇大小，与 过渡的半宽。其他研究将包括 麻醉药对阿司匹林作用下过渡动力学的影响 新型体积微扰动态量热计及其对环境的影响 脂类相关蛋白荧光噪声。最重要的一个方面 这项研究的重点将是调查 麻醉药对胰腺磷脂酶A2激活的影响 我们提出了一个定量描述数据的模型 脂肪酶在磷脂酰胆碱囊泡底物上的作用。 这个模型有三个关键成分：酶结合 优先于凝胶状态的脂类；激活 过程包括在膜表面形成二聚体；以及 激活率与大小成正比 膜内的结构波动。后一个方面 该模型在开发一个 自ALL以来对酶调节机制的认识 膜成分可能会改变膜的波动。 拟议的一系列实验将提供一个数据库 几个热力学和动力学参数与 脂酶系统。这个信息库将允许 具体数量陈述的构建，描述 麻醉作用与其对膜结构的影响 以及膜介导的蛋白质功能。 反应微量热法、差示扫描量热法和 本次调查将使用PH值统计技术。这 实验室在这些技术方面拥有不同寻常的专业知识 将得到其他程序的补充，如荧光 测量、电子显微镜和核磁共振。 实验工作将得到计算机模拟的补充 (例如，蒙特卡罗计算)描述复合材料的模型 实验结果的一部分。

It is our hypothesis that membrane structural fluctuations play an important role in the modulation of biological function. Assuming that the site of anesthetic action is the lipid matrix of the biological membrane, we propose that the initial event in anesthesia is alteration of such fluctuations. To test this hypothesis we plan to use the gel-liquid crystalline transition of single bilayer vesicles as a model for such fluctuations and investigate the affect of various anesthetics on the melting temperature, and the average cluster size which is related to the half-width of the transition. Additional studies will include the effect of anesthetics on the kinetics of the transition using a novel volume perturbation, dynamic calorimeter and their effect on lipid-related protein fluorescence noise. A most important aspect of this study will be the investigation of the effect of anesthetics on the activation of pancreatic phospholipase A2 for which we have proposed a model which quantitatively describes data for the lipase's action on phosphatydilcholine vesicle substrates. This model has three key ingredients: the enzyme binds preferentially to the lipid in the gel state; the activation process involves dimer formation on the membrane surface; and the rate of activation is directly proportional to the magnitude of structural fluctuations within the membrane. This latter aspect of the model is particularly important in developing an understanding of mechanisms of enzyme regulation since all membrane constituents can potentially alter membrane fluctuations. The proposed series of experiments will provide a library of data of several thermodynamic and kinetic parameters related to the lipid-enzyme system. This library of information will allow construction of specific quantitative statements describing anesthetic action in terms of its influence on membrane structure and membrane-mediated protein function. Reaction microcalorimetry, differential scanning calorimetry and pH stat technique will be used in this investigation. This laboratory possesses unusual expertise in these techniques which will be complemented by other procedures such as fluorescence measurements, electron microscopy and nuclear magnetic resonance. The experimental worked will be supplemented by computer simulation (e.g. Monte Carlo Calculations) of models describing the composite of the experimental results.