Sendra Véronique, Cannella Dominique, Bersch Beate, Fieschi Franck, Ménage Stéphane, Lascoux David, Covès Jacques
Laboratoire des Protéines Membranaires, Institut de Biologie Structurale-Jean-Pierre Ebel, UMR 5075 CNRS-CEA-UJF, 41, rue Jules Horowitz, 38027 Grenoble Cedex, France.
Biochemistry. 2006 May 2;45(17):5557-66. doi: 10.1021/bi060328q.
The copH gene is one of the 19 open reading frames (ORFs) found in the cop cluster borne by the large plasmid pMol30 in Cupriavidus metallidurans CH34. The entire cluster is involved in detoxification of copper from the cytoplasm as well as from the periplasm. The function of the corresponding protein, CopH, is not yet clear, but it seems to be involved in the late response phase. We have cloned copH and overproduced and purified the corresponding protein. CopH is rather unique as only one paralog can be found in the databases. It is a dimeric protein with a molecular mass of 13 200 Da per subunit and located in the periplasm. The metal binding properties of CopH were examined by using a series of techniques such as UV-visible spectroscopy, circular dichroism (CD), electron paramagnetic resonance (EPR), surface plasmon resonance (SPR), mass spectrometry, and nuclear magnetic resonance (NMR). All together, the corresponding data are consistent with a dimeric protein containing one metal-binding site per subunit. These sites have a high affinity for Cu(II) but can also bind zinc or nickel. CopH does not contain any cysteines or methionines but contains two histidines. EPR and UV-visible features are consistent with the presence of Cu(II) type 2 centers in a nitrogen ligand field. SPR data confirm the involvement of the histidine residues in copper binding. CD and NMR data reveal that CopH is partially unfolded.
copH基因是在嗜金属贪铜菌CH34的大质粒pMol30携带的cop基因簇中发现的19个开放阅读框（ORF）之一。整个基因簇参与从细胞质以及周质中去除铜的解毒过程。相应蛋白质CopH的功能尚不清楚，但它似乎参与后期反应阶段。我们已经克隆了copH并过量表达和纯化了相应的蛋白质。CopH相当独特，因为在数据库中只能找到一个旁系同源物。它是一种二聚体蛋白质，每个亚基的分子量为13200 Da，位于周质中。通过使用一系列技术，如紫外可见光谱、圆二色性（CD）、电子顺磁共振（EPR）、表面等离子体共振（SPR）、质谱和核磁共振（NMR），研究了CopH的金属结合特性。总的来说，相应的数据与每个亚基含有一个金属结合位点的二聚体蛋白质一致。这些位点对Cu（II）具有高亲和力，但也可以结合锌或镍。CopH不包含任何半胱氨酸或甲硫氨酸，但包含两个组氨酸。EPR和紫外可见特征与氮配体场中2型Cu（II）中心的存在一致。SPR数据证实组氨酸残基参与铜结合。CD和NMR数据表明CopH部分展开。
