MULTINUCLEAR NMR PROBES OF BIOLOGICAL SYSTEMS

总体目标是开发和应用最先进的核磁共振技术 的溶液结构性质的测定技术 复杂的生物大分子，以阐明其机制， 行动上 这些研究所要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 亲环素 金属硫蛋白（MT）：将使用各种NMR方法来充分 解决这两个结构和金属离子结合性能， 普遍存在的、金属离子诱导的、低分子量的、富含半胱氨酸的 金属结合蛋白从各种物种中分离出来， 进化史（真菌、酵母、无脊椎动物和哺乳动物）。 这些 这些特性保证了这种蛋白质在生物学中的最终重要性。 细胞对必需金属（锌、铜）和非必需金属（镉、汞）的调节 离子。 因此，在这些研究中将尽一切努力阐明 这些不同MT的结构的那些方面改变了其 金属离子结合性质并将该数据与假定的， 但尚未明确的功能。 亲环素是一种普遍存在的低分子量（17 Kd）胞浆蛋白 具有高特异性结合环孢菌素A，一种有效的免疫抑制剂 临床上用于预防肾、肝和心脏移植 排斥反应 具体目标将是使用一维和二维 1H NMR方法来阐明溶液的结构特性 亲环素，特别是当它们涉及亲环素的结合位点时， 环孢素。 像这样的研究对于 了解这种蛋白质在药物活性中的作用， 环孢菌素及其活性代谢物，以及结构-活性 环孢菌素的关系。 长期目标是利用 核磁共振，结合其他生物化学方法，以表征 亲环素与其内在天然配体的复合物，当后者是 与世隔绝 这些研究将提供重要的新见解， 环孢菌素作用的分子机制及其潜在的 亲环素在细胞凋亡调控中的生理意义 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.