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STRUCTURE AND REACTIVITY OF METALLOTHIONEIN

金属硫蛋白的结构和反应性

基本信息

批准号：
3277247
负责人：
DAVID Harold PETERING
金额：
$ 2.77万
依托单位：
UNIVERSITY OF WISCONSIN MILWAUKEE
依托单位国家：
美国
项目类别：
财政年份：
1982
资助国家：
美国
起止时间：
1982-04-01 至 1986-03-31
项目状态：
已结题

项目摘要

This proposal describes a series of investigations designed to elucidate the coordination environments of the multiple metal ions in metallothionein and characterize their recently recognized chemical reactivities. Since this sulfhydryl-rich metalbinding protein is a normal cellular constituent in humans and virtually all other organisms and can be induced by the administration of heavy metals, including cadmium, and by a variety of physiological stresses, metallothionein is believed to play an important role in zinc and copper metabolism, heavy metal detoxification, and organismic stress response. A variety of physicochemical techniques will be utilized to elucidate detailed structure-function relationships. Our principal structural tool will be 113Cd NMR, which is a uniquely powerful probe of metallothionein structure and which was recently used to prove that cadmium is grouped in two discrete metalthiolate clusters in the protein. The generality of the two-cluster arrangement will be tested by investigating the 113Cd NMR parameters of metallothioneins from several diverse sources. In concert with 13C NMR studies, the coordination spheres of both cadmium and zinc will be characterized under a variety of conditions. Detailed structural, kinetic and thermodynamic investigations of the formation of metallothionein from metal ions and the apo-protein, thionein, will provide insights into the possible mechanism of its in vivo assembly. Kinetic and mechanistic studies of metal exchange between thionein-bound zinc and cadmium, and the free ions will be carried out, with parallel 113Cd NMR investigation of the differential reactivities of the clusters to metal exchange. Novel reactions of metallothionein including zinc transfer to apo-carbonic anhydrase, EDTA extraction of the metal ions, and disulfide interchange reaction of the thiolates with DTNB will be reinvestigated to determine the relationship of the metal clusters on the reaction kinetics. Novel cadmium complexes with assymetric dithiol chelating ligands will be synthesized as model compounds for 113NMR studies.

该提案描述了一系列旨在阐明金属硫蛋白中多种金属离子的配位环境并表征它们最近被认可的化学反应性。以来这种富含巯基的金属结合蛋白是一种正常的细胞成分，在人类和几乎所有其他生物体中，重金属的管理，包括镉，并通过各种在生理应激中，金属硫蛋白被认为在在锌和铜的代谢，重金属解毒，器官应激反应各种物理化学技术将用于阐明详细的结构-功能关系。我们主要的结构工具将是113 Cd NMR，这是一种独特的强大的金属硫蛋白结构的探针，最近被用来证明镉被分组在两个离散的金属硫醇盐簇中，蛋白两个集群安排的通用性将通过研究了几种金属硫蛋白的113 Cd NMR参数，不同的来源。与13 C NMR研究一致，配位球镉和锌的特性将在各种条件详细的结构、动力学和热力学研究从金属离子和脱辅基蛋白形成金属硫蛋白，硫蛋白，将提供其在体内的可能机制的见解组装件. 金属离子交换的动力学和机理研究硫蛋白结合的锌和镉，和自由离子将进行，用平行113 Cd NMR研究了金属离子交换。金属硫蛋白的新反应包括锌转移到脱碳酸酐酶，EDTA提取的金属离子以及硫醇盐与DTNB的二硫键交换反应将重新研究，以确定金属簇的关系关于反应动力学。新型不对称二硫醇镉配合物螯合配体将被合成为用于113 NMR研究的模型化合物。