MULTINUCLEAR NMR PROBES OF BIOLOGICAL SYSTEMS

长期目标是开发和应用最先进的核磁共振技术 一种溶液结构性质的测定技术 参与金属动态平衡的蛋白质无处不在， 金属硫蛋白，MTS。这一信息对于澄清是必不可少的 金属硫蛋白在金属调节中作用的分子机制。一个 拟议研究的组成部分是为 用于大分子研究的新核磁共振新方法的适应/改进。 虽然详细的结构信息最近可以通过 含镉、锌哺乳动物的核磁共振和X射线结晶学方法 MT，含铜的化合物没有类似的结构信息 哺乳动物MT的形式，也不是来自其他物种的MTS (酿酒酵母、粗糙脉孢菌)。特殊的核磁共振实验 提出了使用银离子作为化学和磁性(自旋)的 I=1/2)同象金属离子替代铜测定痕量铜 这种金属形式的MT的溶液结构。还将使用核磁共振方法 用于海洋MTS全溶液结构的测定 甲壳类、美国龙虾和蓝蟹，它们捆绑着 二价金属离子，镉和锌，和/或一价金属离子，铜。 从这些来源中选择的MT样本，其中将包括重组 中国仓鼠MT的形式，将同时和/或随后使用 在其目标是描绘动力学和 与该蛋白的生理作用相关的机制特征 金属动态平衡。 人们很容易猜测，由于金属的类比- 转录因子和MT中的结合基序，后者 可能在细胞修复过程中起到金属调节作用， 生长和分化。

The long-term objectives are to develop and apply state-of-the-art NMR techniques to the determination of the solution structural properties of a ubiquitously distributed protein involved in metal homeostasis, metallothioneins, MTs. This information is essential for the elucidation of the molecular mechanism of action of MT in metal regulation. An integral component of the proposed research is to contribute the adaptation/refinement of novel new NMR methods for macromolecular studies. While detailed structural information has recently become available through NMR and X-ray crystallographic methods for the Cd, Zn containing mammalian MT, similar structural information is not available for the Cu+ containing forms of the mammalian MT, nor for the MTs from other species (Saccharomyces cerevisiae, Neurospora crassa). Specific NMR experiments are proposed using the Ag+ ion as a chemically and magnetically (spin I=1/2) isomorphic metal ion substitute for Cu+ for the determination of the solution structure of this metal form of MT. NMR methods will also be used for the full solution structure determination of MTs from the marine crustaceans, the American lobster and the blue crab, which bind the divalent metal ions, Cd and Zn, and/or the monovalent metal ion, Cu. Selected MT samples from these sources, which will include recombinant forms of Chinese hamster MT, will concurrently and/or subsequently be used in experiments whose objectives are to delineate the kinetics and mechanistic features associated with this protein's physiological role in metal homeostasis. It is tempting to speculate that because of the analogies of the metal- binding motifs in both the transcription factors and MT, that the latter could have a metalloregulatory function in cellular repair processes, growth, and differentiation.