CHAPERONE TARGET INTERACTIONS

铜是真核生物的必需元素。由于中等电位下氧化还原化学的简便性，它 许多关键酶的活性部位的功能涉及广泛的各种过程，包括 神经递质和神经肽激素的生物合成、生物胺代谢与结缔组织 形成、色素沉着、呼吸和抗氧化防御。新的发现将某些癌症与 铜介导的酶活性，例如观察到铜酶PAM在 内分泌肿瘤是肿瘤生长的中介。另一个耐人寻味的癌症治疗的重要环节是 CTR1高亲和力铜转运体在顺铂摄取细胞中的募集 未知的机制。由于铜既是必需的，也是潜在的有毒物质，细胞进化得很复杂 分子机器将水平维持在一个狭窄的浓度范围内，并未能实现 脂代谢抑制导致与铜缺乏或过量有关的疾病，包括与遗传有关的疾病 铜转运酶ATP7A(孟克斯病)和ATP7B(威尔逊病)突变。 铜稳态异常的重要后果也与神经退行性变有关 阿尔茨海默病患者淀粉样斑块形成的病理研究。因此，对铜的研究 摄取、运输和贩运对人类健康非常重要。在真核生物中，铜是通过 膜结合的高亲和力转运体CTR1，并通过 金属配位体穿梭。铜稳态的一个重要原则是假设CTR1将 铜被输送到伴侣，而伴侣又将金属运送到其作用部位。然而，两国之间的互动 带有伴侣的CTR1还没有得到实验验证。因此，PPG的项目2将利用 HCTR1结构生物学研究的最新进展：(I)铜与CTR1的结合方式；(Ii) CTR1与伴侣HCCS的相互作用，以及(Iii)伴侣和ATPase的作用机制 与它们的目标铜酶相互作用并将铜转移到它们的目标铜酶。硒标记联用技术 X射线吸收光谱学(在上一次赠款期间开发的)将在这些研究中广泛使用。

Copper is an essential element in eukaryotic organisms. Due to facile redox chemistry at moderate potentials, it functions at the active sites of many critical enzymes involved in a wide variety of processes, including neurotransmitter and neuropeptide hormone biosynthesis, biogenic amine metabolism and connective tissue formation, pigmentation, respiration, and antioxidant defense. New findings have linked certain cancers to copper mediated enzyme activity, for example the observation that the expression of the cuproenzyme PAM in endocrine tumors mediates tumor growth. Another intriguing link of importance to cancer therapy, is the recruitment of the CTR1 high affinity copper transporter in the uptake of cis-platin into cells by a presently unknown mechanism. Because copper is both essential and potentially toxic, cells have evolved complex molecular machinery to maintain levels within a narrow range of concentration, and failure to achieve liomeostasis results in diseases related to either Cu deficiency or excess which include those involving genetic mutations in the copper transporting ATPases, ATP7A (Menkes disease) and ATP7B (Wilson disease). Important consequences of aberrant copper homeostasis have also been implicated in neurodegenerative disorders, and in the pathology of amyloid plaque formation in Alzheimers. Therefore, the study of copper uptake, transport and trafficking is of great importance to human health. In eukaryotes, copper is imported by the membrane associated high affinity transporter CTR1, and thence distributed to target proteins via metallochaperone shuttles. An important tenet of copper homeostasis is the hypothesis that CTR1 hands off copper to the chaperones which in turn transport the metal to its sites of action. However, the interaction of CTR1 with chaperones has not yet been experimentally verified. Project 2 of the PPG will therefore leverage recent advances in the structural biology of hCTRl to investigate (i) the mode of copper binding to CTR1, (ii) the interaction of CTR1 with the chaperone hCCS, and (iii) the mechanisms by which chaperones and ATPases interact with and transfer copper to their target cuproenzymes. The technique of selenium-labeling coupled to X-ray absorption spectroscopy (developed in the last grant period) will be used extensively in these studies.