Ricci G, Del Boccio G, Pennelli A, Lo Bello M, Petruzzelli R, Caccuri A M, Barra D, Federici G
Institute of Biochemical Sciences, University of Chieti G. D'Annunzio, Italy.
J Biol Chem. 1991 Nov 15;266(32):21409-15.
Human placenta glutathione transferase (EC 2.5.1.18) pi undergoes an oxidative inactivation which leads to the formation of an inactive enzymatic form which is homogeneous in several chromatographic and electrophoretic conditions. This process is pH dependent, and it occurs at appreciable rate in alkaline conditions and in the presence of metal ions. Dithiothreitol treatment completely restores the active form. -SH titration data and electrophoretic studies performed both on the oxidized and reduced forms indicate that one intrachain disulfide is formed, probably between the two faster reacting cysteinyl groups of each subunit. By the use of a specific fluorescent thiol reagent the disulfide forming cysteines have been identified as the 47th and 101th residues. The disulfide formation causes changes in the tertiary structure of this transferase as appears by CD, UV, and fluorometric analyses; evidences are provided that one or both tryptophanyl residues of each subunit together with a number of tyrosyl residues are exposed to a more hydrophilic environment in the oxidized form. Moreover, electrophoretic data indicate that the subunit of the oxidized enzyme has an apparent molecular mass lower than that of the reduced transferase, thereby confirming structural differences between these forms.
人胎盘谷胱甘肽转移酶（EC 2.5.1.18）π经历氧化失活，导致形成一种在多种色谱和电泳条件下均一的无活性酶形式。此过程依赖于pH值，在碱性条件下且存在金属离子时以可观的速率发生。二硫苏糖醇处理可完全恢复活性形式。对氧化型和还原型进行的-SH滴定数据和电泳研究表明，形成了一个链内二硫键，可能在每个亚基的两个反应较快的半胱氨酰基团之间。通过使用特定的荧光硫醇试剂，已确定形成二硫键的半胱氨酸为第47位和第101位残基。如圆二色性（CD）、紫外（UV）和荧光分析所示，二硫键的形成导致该转移酶的三级结构发生变化；有证据表明，每个亚基的一个或两个色氨酸残基以及一些酪氨酸残基在氧化形式下暴露于更亲水的环境中。此外，电泳数据表明，氧化酶的亚基表观分子量低于还原型转移酶，从而证实了这些形式之间的结构差异。
