MULTINUCLEAR NMR PROBES OF BIOLOGICAL SYSTEMS

总体目标是开发和应用最先进的核磁共振 测定溶液结构性质的技术 复杂的生物大分子，以阐明其机制 行动。 这些将研究的特定生物系统 方法是金属硫蛋白和环孢菌素结合蛋白，称为 亲环蛋白。 金属硫蛋白（MT）：将使用多种NMR方法来充分 解决了这种结构和金属离子结合特性 普遍存在、金属离子诱导、低分子量、富含半胱氨酸 从各种物种中分离出来的金属结合蛋白，其长期 进化史（真菌、酵母、无脊椎动物和哺乳动物）。 这些 正是这种特性保证了这种蛋白质在生物体内的最终重要性。 必需金属（锌、铜）和非必需金属（镉、汞）的细胞调节 离子。 因此，这些研究将尽一切努力来阐明 这些不同MT结构的那些方面改变了它的 金属离子结合特性并将该数据与假设相关联， 但该蛋白质家族的功能尚未明确。 亲环蛋白是一种普遍存在的低分子量 (17 Kd) 胞质蛋白 对结合环孢菌素 A（一种有效的免疫抑制剂）具有高度特异性 临床用于预防肾、肝、心脏同种异体移植 拒绝。 具体目标是使用一维和二维 1H NMR 方法阐明溶液结构特性 亲环蛋白，特别是因为它们与结合位点有关 环孢菌素。 诸如此类的研究对于 了解该蛋白质在药理活性中的作用 环孢菌素及其活性代谢物及其结构活性 环孢菌素的关系。 长期目标是使用 NMR 与其他生化方法结合，可表征 当后者是亲环蛋白与其内在天然配体的复合物时 孤立的。 这些研究将为我们提供重要的新见解 环孢菌素的分子作用机制及其潜力 亲环蛋白在细胞调节中的生理意义 新陈代谢。

The overall objectives are to develop and apply state-of-the-art NMR techniques to the determination of the solution structural properties of complex biological macromolecules in order to elucidate their mechanism of action. The specific biological systems that will be studied by these methods are metallothioneins and a cyclosporin binding protein, termed cyclophilin. Metallothioneins (MTs): A variety of NMR methods will be used to fully resolve both the structural and metal-ion binding properties of this ubiquitous, metal-ion inducible, low molecular weight, cysteine-rich metal-binding protein isolated from various species in its long evolutionary history (fungus, yeast, invertebrate and mammalian). These very properties assures the ultimate importance of this protein in the cellular regulation of essential (Zn, Cu) and nonessential (Cd, Hg) metal ions. As a result, every effort will be made in these studies to elucidate those aspects of the structure of these different MTs which alter its metal-ion binding properties and to correlate this data with the presumed, but as yet undefined, function of this family of proteins. Cyclophilin is a ubiquitous, low molecular weight (17 Kd) cytosolic protein with high specificity for binding cyclosporin A, a potent immunosuppressant used clinically for the prevention of kidney, liver and heart allograft rejection. The specific objectives will be to use one and two-dimensional 1H NMR methods to elucidate the solution structural properties of cyclophilin, particularly as they relate to the binding site for cyclosporin. Studies such as these are of utmost importance to understanding the role of this protein in the pharmacologic activity of cyclosporin and its active metabolites, and the structure-activity relationships for cyclosporin. The long-term objectives would be to use NMR, in conjunction with other biochemical methods, to characterize the complex of cyclophilin with its intrinsic natural ligand when the latter is isolated. These studies would provide important new insight into the molecular mechanism of action of cyclosporin and the potential physiological significance of cyclophilin in the regulation of cellular metabolism.