Heterodimer formation between superoxide dismutase and its copper chaperone
Heterodimer formation between superoxide dismutase and its copper chaperone
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DOI：
10.1021/bi002207a
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发表时间：
2000-12-05
期刊：
影响因子：
2.9
通讯作者：
Rosenzweig, AC
中科院分区：
文献类型：
--
作者：
Lamb, AL;Torres, AS;Rosenzweig, AC
铜锌超氧化物歧化酶(SOD1)在体内被铜超氧化物歧化酶(CCS)的伴侣蛋白激活。CCS识别并与SOD1对接以插入金属离子的分子机制尚不清楚。提出了铜转移过程中齐聚状态的两种模型：一种是由一种CCS单体和一种SOD1单体组成的杂二聚体，另一种是包含两个同源二聚体相互作用的二聚体。我们研究了野生型SOD_1(WtSOD_1)和铜配体His 48被苯丙氨酸取代的突变型SOD_1(H48F-SOD_1)的载铜酵母CCS(YCCS)和酵母SOD_1之间蛋白质-蛋白质复合体的形成。根据凝胶过滤层析、动态光散射、分析超速离心法和化学交联实验，YCCS和突变体SOD1形成了杂二聚体的正确分子质量的复合体。未检测到高阶齐聚物。锌的存在促进了杂二聚体的形成，但不依赖于YCCS的载铜量。与wtSOD1形成的络合物相比，与H38F-SOD1形成的络合物更稳定，说明后者是一种更具暂时性的物种。值得注意的是，只有在锌存在的情况下，载铜YCCS和wtSOD1之间形成的异二聚体才伴随着SOD1的激活。这些发现与结构、生化和遗传学研究结合在一起，有力地表明在体内酵母SOD1的铜负载是通过异源二聚体发生的。
Copper, zinc superoxide dismutase (SOD1) is activated in vivo by the copper chaperone for superoxide dismutase (CCS). The molecular mechanisms by which CCS recognizes and docks with SOD1 for metal ion insertion are not well understood. Two models for the oligomerization state during copper transfer have been proposed: a heterodimer comprising one monomer of CCS and one monomer of SOD1 and a dimer of dimers involving interactions between the two homodimers. We have investigated protein-protein complex formation between copper-loaded and apo yeast CCS (yCCS) and yeast SOD1 for both wild-type SOD1 (wtSOD1) and a mutant SOD1 in which copper ligand His 48 has been replaced with phenylalanine (H48F-SOD1). According to gel filtration chromatography, dynamic light scattering, analytical ultracentrifugation, and chemical cross-linking experiments, yCCS and this mutant SOD1 form a complex with the correct molecular mass for a heterodimer. No higher order oligomers were detected. Heterodimer formation is facilitated by the presence of zinc but does not depend on copper loading of yCCS. The complex formed with H38F-SOD1 is more stable than that formed with wtSOD1, suggesting that the latter is a more transient species. Notably, heterodimer formation between copper-loaded yCCS and wtSOD1 is accompanied by SOD1 activation only in the presence of zinc. These findings, taken together with structural, biochemical, and genetic studies, strongly suggest that in vivo copper loading of yeast SOD1 occurs via a heterodimeric intermediate.