Sarkar B
Department of Structural Biology and Biochemistry, The Hospital for Sick Children, Toronto, Ont., Canada.
J Inorg Biochem. 2000 Apr;79(1-4):187-91. doi: 10.1016/s0162-0134(99)00249-4.
Copper is an essential trace element which forms an integral component of many enzymes. While trace amounts of copper are needed to sustain life, excess copper is extremely toxic. An attempt is made here to present the current understanding of the normal transport of copper in relation to the absorption, intracellular transport and toxicity. Wilson disease is a genetic disorder of copper transport resulting in the accumulation of copper in organs such as liver and brain which leads to progressive hepatic and neurological damage. The gene responsible for Wilson disease (ATP7B) is predicted to encode a putative copper-transporting P-type ATPase. An important feature of this ATPase is the presence of a large N-terminal domain that contains six repeats of a copper-binding motif which is thought to be responsible for binding this metal prior to its transport across the membrane. We have cloned, expressed and purified the N-terminal domain (approximately 70 kD) of Wilson disease ATPase. Metal-binding properties of the domain showed the protein to bind several metals besides copper; however, copper has a higher affinity for the domain. The copper is bound to the domain in Cu(I) form with a copper: protein ratio of 6.5:1. X-ray absorption studies strongly suggest Cu(I) atoms are ligated to cysteine residues. Circular dichroism spectral analyses suggest both secondary and tertiary structural changes upon copper binding to the domain. Copper-binding studies suggest some degree of cooperativity in binding of copper. These studies as well as detailed structural information of the copper-binding domain will be crucial in determining the specific role played by the copper-transporting ATPase in the homeostatic control of copper in the body and how the transport of copper is interrupted by mutations in the ATPase gene.
铜是一种必需的微量元素，是许多酶的重要组成部分。维持生命需要微量的铜，但过量的铜具有极强的毒性。本文旨在阐述目前对铜的正常转运过程的理解，包括其吸收、细胞内转运及毒性。威尔逊病是一种铜转运的遗传性疾病，会导致铜在肝脏和大脑等器官中蓄积，进而引发进行性肝脏和神经损伤。导致威尔逊病的基因（ATP7B）预计编码一种假定的铜转运P型ATP酶。该ATP酶的一个重要特征是存在一个大的N端结构域，其中包含六个铜结合基序重复序列，据认为这些序列在铜跨膜转运之前负责结合这种金属。我们已经克隆、表达并纯化了威尔逊病ATP酶的N端结构域（约70 kD）。该结构域的金属结合特性表明，除了铜之外，该蛋白还能结合多种金属；然而，铜对该结构域具有更高的亲和力。铜以Cu(I)形式与该结构域结合，铜与蛋白质的比例为6.5:1。X射线吸收研究强烈表明，Cu(I)原子与半胱氨酸残基相连。圆二色光谱分析表明，铜与该结构域结合后，二级和三级结构均发生变化。铜结合研究表明，铜结合存在一定程度的协同作用。这些研究以及铜结合结构域的详细结构信息对于确定铜转运ATP酶在体内铜稳态控制中所起的具体作用，以及ATP酶基因突变如何中断铜的转运至关重要。
