Physiological roles of a new copper-binding protein : relationship between new copper-binding protein and hereditary copper

铜是一种必需的微量元素，需要在必要的浓度和毒性之间保持微妙的细胞平衡。金属硫蛋白（MT）是一种低分子量的重结合蛋白，在铜的体内平衡和解毒中起着重要作用。本研究采用一种新的铜结合蛋白和铜-金属硫蛋白的分离纯化方法，将单次或多次注射铜诱导的低分子量铜结合蛋白鉴定为铜-金属硫蛋白和/或一种新的铜结合蛋白。我们通过证明一种新的铜诱导蛋白的存在解决了这一混乱，该蛋白不属于MT组，因为该蛋白是低半胱氨酸含量的，并且每个分子结合两个铜原子。结果表明，在大鼠肝脏中，单次注射Cu仅诱导了Cu-MT，多次注射Cu则同时诱导了Cu-MT和新蛋白的表达。这一结果表明，当肝脏Cu含量增加并达到一定的阈值时，Cu-MT不能充分解毒的过量Cu合成了新的蛋白质，从而起到解毒Cu的作用。此外，我们确定了Menke病（X连锁Cu代谢障碍）模型Macular小鼠肾脏中过量Cu为Cu-MT。皮质近曲小管细胞中铜金属硫蛋白占优势。MT mRNA也在皮层中观察到，表明该蛋白质在该区域生物合成。另一方面，新的铜结合蛋白在黄斑小鼠的肾脏中几乎没有检测到。根据这些结果，新蛋白被认为在铜代谢中起关键作用。

Cu is an essential trace element which requires a delicate cellular balance between a necessary concentration and toxicity. Metallothionein (MT), a low molecular weight heavy-binding protein, plays an important role in Cu homeostasis and detoxification. In this study, the Cu-binding low molecular weight proteins induced by single or multiple Cu-injection rats were identified as Cu-MTs and/or a new Cu-binding protein using a new purification procedure of both of the new protein and Cu-MT under the aerobic condition.There were many debate on the existence of MT and other Cu-binding proteins which appeared in tissues of Cu-loaded animals. We settled the confusion by demonstrating of the presence of a new Cu-induced protein which did not belong to MT group, because the protein was a low cystein content and bound two Cu atoms per a molecule. We did show that only Cu-MT was induced by a single injection of Cu in rat liver and both of Cu-MT and the new protein appeared by the multiple injections of Cu. This result suggests that new protein was synthesized by excessive Cu which was not sufficiently detoxified by Cu-MT to serve detoxification of Cu when the hepatic Cu content increased and reached a certain threshold value.In addition, we established that excess amounts of Cu in the kidney of Macular mice, a model for Menke's disease (X-linked disorder of Cu metabolism), were found as Cu-MT. The Cu-MT was predominant in the proximal convoluted tubule cells of the cortex. MT mRNA was also observed in the cortex, indicating that the protein was biosynthesized in this region. On the other hand, the new Cu-binding protein was hardly detected in the kidney of macular mice. From these results the new protein was considered to play a key role in Cu-metabolism.