Spatial proximity of Cys(113), Cys(172), and Cys(422) in the metalloactivation domain of the ArsA ATPase
Spatial proximity of Cys(113), Cys(172), and Cys(422) in the metalloactivation domain of the ArsA ATPase
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DOI：
10.1074/jbc.271.40.24465
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发表时间：
1996-10-04
影响因子：
4.8
通讯作者：
Rosen, BP
中科院分区：
文献类型：
--
作者：
Bhattacharjee, H;Rosen, BP
ArsA ATP酶活性被半金属砷或锑的盐变构激活。激活与ArsA中三个半胱氨酸残基的存在相关：Cys（113），Cys（172）和Cys（422）。为了确定半胱氨酸残基之间的距离，用双功能烷基化剂二溴亚胺处理半胱氨酸取代为丝氨酸的野生型ArsA和ArsA蛋白质，该试剂与彼此相距3-6埃的巯基对反应，形成荧光加合物。其中单个半胱氨酸残基通过定点诱变改变的ArsA蛋白仍然形成荧光加合物。其中三个半胱氨酸残基中的两个被取代的蛋白质不形成荧光加合物。这些结果表明Cys（113）、Cys（172）和Cys（422）彼此非常接近。我们提出了一个模型，其中As（III）或Sb（III）与这三个半胱氨酸在三角锥几何形状的相互作用，形成一个新的软金属硫醇笼。
ArsA ATPase activity is allosterically activated by salts of the semimetal arsenic or antimony. Activation is associated with the presence of three cysteine residues in ArsA: Cys(113), Cys(172), and Cys(422). To determine the distance between cysteine residues, wild type ArsA and ArsA proteins with cysteine to serine substitutions were treated with the bifunctional alkylating agent dibromobimane, which reacts with thiol pairs within 3-6 Angstrom of each other to form a fluorescent adduct. ArsA proteins in which single cysteine residues were altered by site-directed mutagenesis still formed fluorescent adducts. Proteins in which two of the three cysteine residues were substituted did not form fluorescent adducts, These results demonstrate that Cys(113), Cys(172), and Cys(422) are in close proximity of each other. We propose a model in which As(III) or Sb(III) interacts with these three cysteines in a trigonal pyramidal geometry, forming a novel soft metal-thiol cage.