Frederick National Laboratory for Cancer Research, Frederick, Maryland 21702, United States.
Chem Res Toxicol. 2012 Dec 17;25(12):2670-7. doi: 10.1021/tx3003142. Epub 2012 Nov 9.
Attachment of glutathione (GSH) to cysteine residues in proteins (S-glutathionylation) is a reversible post-translational modification that can profoundly alter protein structure and function. Often serving in a protective role, for example, by temporarily saving protein thiols from irreversible oxidation and inactivation, glutathionylation can be identified and semiquantitatively assessed using anti-GSH antibodies, thought to be specific for recognition of the S-glutathionylation modification. Here, we describe an alternate mechanism of protein glutathionylation in which the sulfur atoms of the GSH and the protein's thiol group are covalently bound via a cross-linking agent, rather than through a disulfide bond. This form of thiol cross-linking has been shown to occur and has been confirmed by mass spectrometry at the solution chemistry level, as well as in experiments documenting the potent antiproliferative activity of the bis-diazeniumdiolate Double JS-K in H1703 cells in vitro and in vivo. The modification is recognized by the anti-GSH antibody as if it were authentic S-glutathionylation, requiring mass spectrometry to distinguish between them.
谷胱甘肽（GSH）与蛋白质半胱氨酸残基的结合（S-谷胱甘肽化）是一种可逆的翻译后修饰，可以深刻改变蛋白质的结构和功能。例如，它通常具有保护作用，通过暂时保存蛋白质巯基免受不可逆氧化和失活，谷胱甘肽化可以使用抗 GSH 抗体进行识别和半定量评估，这些抗体被认为是特异性识别 S-谷胱甘肽化修饰的。在这里，我们描述了一种蛋白质谷胱甘肽化的替代机制，其中 GSH 的硫原子和蛋白质的巯基通过交联剂共价结合，而不是通过二硫键。这种形式的硫交联已被证明在溶液化学水平以及在实验中发生，并通过质谱证实，实验记录了双氮烯二醇 Double JS-K 在体外和体内 H1703 细胞中的强大抗增殖活性。该修饰被抗 GSH 抗体识别为真实的 S-谷胱甘肽化，需要通过质谱来区分它们。
