ANESTHETIC ALTERATION OF PROTEIN LIPID INTERACTIONS

蛋白质脂质相互作用的麻醉改变

• 批准号: 3293134
• 负责人: RODNEY L BILTONEN
• 金额: $ 22.84万
• 依托单位: UNIVERSITY OF VIRGINIA
• 依托单位国家: 美国
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-02-01 至 1995-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3293134
• 关键词: anesthetics; calorimetry; chemical binding; chemical kinetics; computer simulation; conformation; electron microscopy; enzyme mechanism; fluorescence spectrometry; gas chromatography mass spectrometry; gel; light scattering; lipid bilayer membrane; lipid structure; liquid crystal; membrane activity; membrane lipids; membrane structure; microcalorimetry; molecular dynamics; nuclear magnetic resonance spectroscopy; phase change; phospholipase A2; temperature; thermodynamics

It is our hypothesis that membrane structural fluctuations play an important role in the modulation of membrane-protein 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. This hypothesis will be tested using the gel to liquid- crystalline transition and compositional-based phase separations in single bilayer vesicles as models for such fluctuations. The effects of general and local anesthetics on the equilibrium properties of these transitions will be investigated using calorimetric and spectroscopic techniques. Particular emphasis will be placed on quantitative evaluation of the anesthetic-induced changes in the melting temperature and shape of the heat capacity function associated with the thermotropic transitions. The former parameter is a measure of the differential solubility of the anesthetic in the gel and liquid-crystalline states; the latter parameter is related to the details (e.g. cluster size) of the distribution of distinct molecular species of the lipids. The dynamics of the transitions will also be investigated using a novel, volume perturbation, dynamic calorimeter. An important aspect of this study is the investigation of how anesthetics alter the vesicle-surface activation of the enzyme phospholipase A2 which appears to be strongly coupled to lipid structural fluctuations. These structural fluctuations depend upon temperature, lipid composition and reaction products which can induce spontaneous activation of the enzyme. This is particularly important since all membrane constituents can potentially alter membrane fluctuations. The proposed experiments will provide a library of thermodynamic and kinetic information related to define lipid-enzyme system, allowing 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, fluorescence spectroscopy, and pH stat techniques will be used in this investigation. This laboratory possesses unusual expertise in these techniques, which will be complemented by other procedures such as electron microscopy and nuclear magnetic resonance. The experimental work will be supplemented by computer simulation (e.g. Monte Carlo calculations) of the models describing the composite of the experimental results.

我们的假设是膜结构的波动起到了 在膜蛋白功能的调节中起着重要作用。假设 麻醉作用的部位是生物体的脂质基质 膜，我们认为麻醉的最初事件是改变 这样的波动。这一假说将用凝胶来验证-- 单晶的晶化转变和基于成分的相分离 双层囊泡作为这种波动的模型。一般的影响 和局麻药对这些转变的平衡性质的影响 将使用量热和光谱技术进行研究。 将特别强调量化评估 麻醉剂引起的熔化温度和热形状的变化 与热致转变相关的容量函数。前者 参数是麻醉剂在体内的不同溶解度的量度。 凝胶态和液晶态；后一个参数与 不同分子分布的细节(如团簇大小) 脂类的种类。过渡的动力也将是 使用一种新型的、体积微扰的动态量热计进行了研究。一个 这项研究的重要方面是麻醉剂如何 改变磷脂酶A2的囊泡表面活性 似乎与脂质结构波动有很强的耦合关系。这些 结构波动取决于温度、脂质成分和 可诱导酶自发激活的反应产物。 这一点尤其重要，因为所有的膜成分都可以 潜在地改变了膜的波动。拟议中的实验将 提供与以下内容相关的热力学和动力学信息库 定义脂质-酶系统，允许构建特定的定量 描述麻醉作用的陈述，其对 膜结构和膜介导的蛋白质功能。 反应微量热法、差示扫描量热法、荧光 光谱分析和pH值统计技术将在本次调查中使用。 这个实验室在这些技术方面拥有不同寻常的专业知识，这将 得到其他程序的补充，如电子显微镜和核 磁共振。这项实验工作将由计算机辅助 模拟(例如，蒙特卡罗计算)描述 综合实验结果。