Biochemical characterization of the Wilson disease protein and functional expression in the yeast Saccharomyces cerevisiae
Biochemical characterization of the Wilson disease protein and functional expression in the yeast Saccharomyces cerevisiae
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DOI：
10.1074/jbc.272.34.21461
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发表时间：
1997-08-22
影响因子：
4.8
通讯作者：
Gitlin, JD
中科院分区：
文献类型：
--
作者：
Hung, IH;Suzuki, M;Gitlin, JD
肝豆状核变性是一种铜代谢紊乱的疾病，其特征是肝硬变和神经元变性，这是由于编码铜转运P型ATPase的基因发生遗传突变所致。针对Wilson蛋白氨基末端制备的多克隆抗血清在HepG2和Caco细胞裂解产物中检测到一个165 kDa的特异性蛋白，进一步分析表明该蛋白是以单链多肽形式合成的，并在稳态条件下定位于反式高尔基体网络。铜浓度的增加导致这种蛋白质快速移动到细胞质的囊泡室。这种铜特定的威尔逊蛋白的细胞重新分布是一个可逆的过程，不依赖于新的蛋白质合成。野生型而不是突变型的Wilson蛋白在酿酒酵母CCC2 Delta菌株中的表达恢复了铜对多铜氧化酶Fet3p的掺入，为威尔逊蛋白对铜的运输提供了直接证据。综上所述，这些数据揭示了铜代谢的细胞机制具有显着的进化保守性，并为调控铜进入真核细胞的分泌途径提供了一个独特的模型。
Wilson disease is a disorder of copper metabolism characterized by hepatic cirrhosis and neuronal degeneration due to inherited mutations in a gene encoding a putative copper-transporting P-type ATPase. Polyclonal antisera generated against the amino terminus of the Wilson protein detected a specific 165-kDa protein in HepG2 and CaCo cell lysates, Further analysis revealed that this protein is synthesized as a single-chain polypeptide and localized to the trans-Golgi network under steady state conditions. An increase in the copper concentration resulted in the rapid movement of this protein to a cytoplasmic vesicular compartment. This copper-specific cellular redistribution of the Wilson protein is a reversible process that occurs independent of a new protein synthesis. Expression of the wild-type but not mutant Wilson protein in the ccc2 Delta strain of Saccharomyces cerevisiae restored copper incorporation into the multicopper oxidase Fet3p, providing direct evidence of copper transport by the Wilson protein. Taken together these data reveal a remarkable evolutionary conservation in the cellular mechanisms of copper metabolism and provide a unique model for the regulation of copper transport into the secretory pathway of eucaryotic cells.