EPR STUDIES OF MOLECULAR MOTION IN MEMBRANE SYSTEMS

这个方案的目标是描述脂肪和蛋白质的运动 生物系统内膜中的成分以及与这种运动有关 为行使职能干杯。具体来说，我们的目标是：a)描述 膜中脂质-蛋白质相互作用和蛋白质-蛋白质结合 在运动方面，相互之间以及与膜之间的“可溶性酶” 膜组件的行为；b)研究调节这种行为的因素 运动；以及c)将膜组件的运动特性与 行使职能。蛋白质组分运动的研究 将强调使用饱和转移EPR(ST-EPR)来测量 在慢时间域中的运动，大分子和 超分子络合物。自旋标签将被合成并用于 在选择性位置贴标签膜组件以进行优化 释义。光谱将通过计算机模拟进行分析，使用 用于计算各向异性运动谱的新算法。 随着数据的获得，理论的应用可以得到测试、改进 和延展。涉及脂质运动的研究将与 核磁共振研究，以便能够分析自旋标记的可靠性。这个 计划将EPR和ST-EPR的一般应用测试和扩展到 对膜组件运动行为的研究。的主要系统 被研究的包括：a)D-β-羟基丁酸脱氢酶，a 需要脂肪的酶；b)骨骼肌肌浆的钙泵 对催化泵送循环进行了仔细的研究， 应该作为将运动特征与具体特征联系起来的原型 功能细节；c)牛心脏线粒体转氢酶， 由能够控制的单个膜多肽亚单位组成 质子力的激发；d)牛的三磷酸腺苷合成酶(F1-Fo) 心脏线粒体；这种换能器能够相互转换 三磷酸腺苷合成或水解的质子推动力。重组 强调了方法的重要性。具有定义的组成和功能的膜可以 做好准备。在这种情况下，各种条件的影响对 可以研究膜的组成、功能和运动特性 并相互关联。

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.