STRUCTURAL BIOLOGY OF COPPER HOMEOSTASIS

铜稳态的结构生物学

• 批准号: 7690563
• 负责人: VINZENZ UNGER
• 金额: $ 27.75万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-15 至 2014-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7690563
• 关键词: ATP phosphohydrolase; Active Sites; Address; Affinity; Alzheimer' s Disease; Amino Acids; Antineoplastic Agents; Area; Behavior; Binding; Binding Sites; Biochemical; Biology; C-terminal; Cell physiology; Cell secretion; Cells; Cisplatin; Complex; Copper; Creutzfeldt-Jakob Syndrome; Crystallization; Cues; Data; Disease; Docking; Electrons; Engineering; Family; Genetic; Glycophorin A; Goals; Homeostasis; Human; Ions; Letters; Light; Link; Mediating; Membrane; Membrane Proteins; Membrane Transport Proteins; Menkes Kinky Hair Syndrome; Metals; Methods; Microscopic; Molecular; Molecular Chaperones; Mus; Parkinson Disease; Physiological; Process; Proteins; Regulation; Resolution; Role; Sampling; Site; Structure; Testing; Wilson disease protein; Work; Yeasts; base; copper-transporting ATPase; crosslink; electron crystallography; genetic regulatory protein; in vivo; insight; metalloenzyme; neuropathology; novel; orexin A receptor; orexin B receptor; particle; reconstruction; response; spatial relationship; structural biology; tool; trafficking; uptake

Genetic and biochemical studies lend strong support to the idea that the CTR-family of membrane proteins mediate cellular copper uptake. Similarly, the roles of copper transporting ATPases and intracellular copper chaperones in copper secretion and transfer of copper ions to the active sites of cuproenzymes have been firmly established. What remains elusive, is how these different components are integrated at the molecular level, largely because the structures of the membrane proteins involved in these processes are unknown. The longterm goal of this project is to understand at a structural level the mechanism by which copper transporting membrane proteins and cellular copper chaperones interact with each other to accomplish directed flow of copper within cells. Work towards Aim 1 capitalizes on our progress in the structure determination of the human high affinity copper transporter hCTR1. The structure suggests that clustering of the C-termini in the hCTR1 trimer creates a binding site for both copper ions and copper chaperones, which would allow for a direct transfer of copper from hCTR1 to the chaperones. Notably, the amino acid residues forming this site are absent in mammalian CTR2, raising the question whether all CTR-proteins use the same mechanism to distribute copper to downstream acceptors. We propose to solve the structure of hCTR2 by electron crystallography to investigate the hypothesis that in hCTR2 an alternate set of metal binding residues can functionally replace the C-terminal chaperone docking site observed in hCTRl Further pursuing the idea of direct CTR:chaperone interactions, the goal of Aim 2 is to trap CTR:chaperone complexes, and to visualize their structure by electron microscopic single particle methods. Finally, the longterm goal for Aim 3 is to solve the structure of the copper transporting ATPase 7B, which is involved in copper secretion. The key aspect of these studies will be to delineate the spatial relationships between the membrane embedded and cytosolic domains that are critical for function and the regulation of the ATPase's complex trafficking behavior.

遗传学和生化研究对膜蛋白的CTR家庭的观念有很大的支持 介导细胞铜摄取。同样，铜运输ATPases和细胞内铜的作用 铜分泌中的伴侣和铜离子向库酶的活性部位的转移已经 牢固确立。仍然难以捉摸的是如何将这些不同的成分集成在分子上 水平，主要是因为参与这些过程的膜蛋白的结构尚不清楚。 该项目的长期目标是在结构层面上了解铜的机制 运输膜蛋白和细胞铜伴侣相互相互作用以完成 铜在细胞内的定向流。 朝着目标1努力利用我们在人类高亲和力的结构确定方面的进步 铜转运蛋白HCTR1。该结构表明C-termini在HCTR1夹子中的聚类 为铜离子和铜伴侣创建一个结合位点，这将允许直接转移 从HCTR1到伴侣的铜。值得注意的是，在 哺乳动物CTR2，提出了一个问题，是否所有CTR蛋白使用使用相同的机制分发 铜到下游受体。我们建议通过电子晶体学求解HCTR2的结构 研究以下假设：在HCTR2中，一组替代的金属结合残基可以在功能上替代 HCTRL观察到的C末端伴侣对接站点进一步追求直接的想法 CTR：伴侣相互作用，AIM 2的目标是捕获CTR：伴侣复合体，并可视化它们 通过电子显微镜单粒子方法结构。最后，目标3的长期目标是解决 运输ATPase 7b的铜的结构，该结构与铜分泌有关。关键方面 这些研究将是描述嵌入膜和胞质之间的空间关系 对功能至关重要的域和ATPase复杂运输行为的调节。