Cu(I) Affinities of the Domain 1 and 3 Sites in the Human Metallochaperone for Cu,Zn-Superoxide Dismutase
Cu(I) Affinities of the Domain 1 and 3 Sites in the Human Metallochaperone for Cu,Zn-Superoxide Dismutase
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DOI：
10.1021/bi201370r
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发表时间：
2012-02-21
期刊：
影响因子：
2.9
通讯作者：
Dennison, Christopher
中科院分区：
文献类型：
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作者：
Allen, Stephen;Badarau, Adriana;Dennison, Christopher
人金属伴侣CCS对铜的传递是激活Cu， zn -超氧化物歧化酶（SOD1）的关键步骤。CCS是一种三结构域蛋白，在结构域1和3中分别与Cu(I)结合CXXC和CXC基序。详细分析了铜与CCS的结合，包括结构域1和3的Cys残基突变为Ser的变体，以及使用单独的结构域1和3结构域，表明CCS能够在这两个结构域中结合1个Cu(I)。在pH 7.5时，结构域1的Cu(I)亲和力约为5 × 10(17) M-1，而结构域3的Cu(I)亲和力至少弱一个数量级。因此，CXXC位点将优先加载Cu(I)，这表明结构域1在金属的获取中起作用。铜通过结构域3传递到靶点，其结构灵活性和在铜传递过程中短暂金属化的能力似乎比CXC基元的Cu(I)亲和力更重要。CCS结构域1的Cu(I)亲和力与另一种细胞质铜金属伴侣蛋白HAH1相当。CCS和HAH1容易交换Cu(I)，提供了在铜运输途径之间发生串扰的机制。
The delivery of copper by the human metallochaperone CCS is a key step in the activation of Cu,Zn-superoxide dismutase (SOD1). CCS is a three-domain protein with Cu(I)-binding CXXC and CXC motifs in domains 1 and 3, respectively. A detailed analysis of the binding of copper to CCS, including variants in which the Cys residues from domains 1 and 3 have been mutated to Ser, and also using separate domain 1 and 3 constructs, demonstrates that CCS is able to bind 1 equiv of Cu(I) in both of these domains. The Cu(I) affinity of domain 1 is approximately 5 X 10(17) M-1 at pH 7.5, while that of domain 3 is at least 1 order of magnitude weaker. The CXXC site will therefore be preferentially loaded with Cu(I), suggesting that domain 1 plays a role in the acquisition of the metal. The delivery of copper to the target occurs via domain 3 whose structural flexibility and ability to be transiently metalated during copper delivery appear to be more important than the Cu(I) affinity of its CXC motif The Cu(I) affinity of domain 1 of CCS is comparable to that of HAH1, another cytosolic copper metallochaperone. CCS and HAH1 readily exchange Cu(I), providing a mechanism whereby cross-talk can occur between copper trafficking pathways.