EPR STUDIES OF MOLECULAR MOTION IN MEMBRANE SYSTEMS

这个建议的目标是描述脂质和蛋白质的运动 生物系统内的膜中的组分，并将这种运动 到行使职能。 具体而言，我们的目标是：a）表征 膜中的脂质-蛋白质相互作用， “可溶性酶”与另一个和膜，在运动方面， B）研究调节这种特性的因素， 运动;以及c）将膜组件的运动特性与 行使职能。 蛋白质运动学研究蛋白质组分运动的学科 将强调使用饱和转移EPR（ST-EPR），它可以测量 在慢时域中的运动，大分子的特性， 超分子复合物 自旋标记将被合成并用于 在选择性位置标记膜组件以优化 解释。 光谱将通过计算机模拟进行分析， 新的可用算法计算各向异性运动谱。 随着数据的获得，理论的应用可以得到测试，改进， 并延长。 涉及脂质运动的研究将与 核磁共振研究，使自旋标记的可靠性可以分析。 的 该计划将测试和扩展EPR和ST-EPR的一般应用， 研究膜的组成部分运动行为的学科。 主要系统， 研究的包括：a）D-β-羟基丁酸脱氢酶， B）骨骼肌肌浆CA++泵 网;催化泵送循环已经仔细研究， 应该作为一个原型，将运动特征与特定 功能细节; c）来自牛心脏线粒体的转氢酶， 由能够控制的单个膜多肽亚基组成， d）来自牛的ATP合成酶（F1-Fo） 心脏线粒体;该换能器能够相互转换 质子动力对ATP合成或水解的影响。 复溶 方法得到强调。 具有确定的组成和功能的膜可以 做好准备 这样，各种条件对 可以研究膜的组成、功能和运动特性 和相关性。

The goal of this proposal is to describe the motion of lipid and protein components in membranes within biological systems and to relate such motion to the exercise of function. Specifically we aim to: a) characterize lipid-protein interactions in membranes, and protein-protein association of "soluble enzymes" with one another and with membranes, in terms of motional behavior of membrane components; b) study factors which modulate such motion; and c) relate motional characteristics of membrane components to the exercise of function. The study of the motion of protein components will emphasize use of satguration transfer EPR (ST-EPR) which can measure motion in the slow time domain, characteristic of macromolecules and supramolecular complexes. Spin labels will be synthesized and used to label membrane components in selective positions to optimize interpretation. The spectra will be analyzed by computer simulation using newly available algorithms for computation of anisotropic motion spectra. As data become available, the application of theory can be tested, refined and extended. Studies involving the motion of lipids will be compared with NMR studies so that the reliability of spin labels can be analyzed. The program will test and extend the general application of EPR and ST-EPR to the study of motional behavior of membrane components. The main systems to be investigated include: a) D-Beta-Hydroxybutyrate dehydrogenase, a lipid-requiring enzyme; b) The CA++ pump of skeletal muscle sarcoplasmic reticulum; the catalytic pumping cycle has been carefully studied and should serve as a prototype to relate motional characteristics to specific functional detail; c) Transhydrogenase from bovine heart mitochondria, consisting of a single membrane polypeptide subunit capable of controlled gneration of protonmotive force; d) ATP synthetase (F1-Fo) from bovine heart mitochonria; this transducer is capable of interconverting protonmotive force to ATP synthesis or hydrolysis. The reconstitution approach is emphasized. Membranes of defined composition and function can be prepared. In this way, the influence of a variety of conditions on membrane composition, function and motional characteristics can be studied and correlated.