MULTINUCLEAR NMR PROBES OF BIOLOGICAL SYSTEMS

总体目标是开发和应用最先进的核磁共振 溶液结构性质的测定技术 复杂的生物大分子以阐明其作用机制 行动。将由这些人研究的特定生物系统 方法是金属硫蛋白和环孢素结合蛋白，称为 亲环素。 金属硫蛋白(MTS)：将充分利用各种核磁共振方法 解决了该化合物的结构和金属离子结合性质 无处不在，金属离子诱导型，低分子量，富含半胱氨酸 从不同物种中分离的金属结合蛋白 进化史(真菌、酵母、无脊椎动物和哺乳动物)。这些 非常性质确保了这种蛋白质在 必需(锌、铜)和非必需(镉、汞)金属的细胞调控 离子。因此，这些研究将尽一切努力澄清 这些不同MT的结构的哪些方面改变了其 金属-离子结合性质并将该数据与推定的， 但到目前为止，这个蛋白质家族的功能还没有确定。 亲环素是一种普遍存在的低分子量(17kD)胞浆蛋白 与一种有效的免疫抑制剂环孢素A结合具有高特异性 临床用于预防同种异体肾、肝、心移植 拒绝。具体目标将是使用一维和二维 用~1H核磁共振方法研究水溶液的结构性质 亲环素，尤其是与结合部位有关的亲环素 环孢素。像这样的研究对 了解该蛋白在血管活性物质药理活性中的作用 环孢素及其活性代谢物及其构效关系 环孢素的关系。长期目标将是使用 核磁共振，结合其他生化方法，以表征 亲环素与其固有天然配体的络合物 与世隔绝。这些研究将为我们提供重要的新见解 环孢素作用的分子机制及其作用潜力 亲环素在细胞调控中的生理学意义 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.