S-Methyl methanethiosulfonate (MMTS) is used in experimental biochemistry for alkylating thiol groups of protein cysteines. Its applications include mainly trapping of natural thiol-disulfide states of redox-sensitive proteins and proteins which have undergone S-nitrosylation. The reagent can also be employed as an inhibitor of enzymatic activity, since nucleophilic cysteine thiolates are commonly present at active sites of various enzymes. The advantage of using MMTS for this purpose is the reversibility of the formation of methylthio mixed disulfides, compared to irreversible alkylation using conventional agents. Additional benefits include good accessibility of MMTS to buried protein cysteines due to its small size and the simplicity of the protection and deprotection procedures. In this study we report examples of MMTS application in experiments involving oxidoreductase (glyceraldehyde-3-phosphate dehydrogenase, GAPDH), redox-regulated protein (recoverin) and cysteine protease (triticain-α). We demonstrate that on the one hand MMTS can modify functional cysteines in the thiol enzyme GAPDH, thereby preventing thiol oxidation and reversibly inhibiting the enzyme, while on the other hand it can protect the redox-sensitive thiol group of recoverin from oxidation and such modification produces no impact on the activity of the protein. Furthermore, using the example of the papain-like enzyme triticain-α, we report a novel application of MMTS as a protector of the primary structure of active cysteine protease during long-term purification and refolding procedures. Based on the data, we propose new lines of MMTS employment in research, pharmaceuticals and biotechnology for reversible switching off of undesirable activity and antioxidant protection of proteins with functional thiol groups.

中文翻译：

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使用S-甲基甲硫基磺酸盐（MMTS）修饰硫醇酶和氧化还原调节蛋白的新应用。

S-甲硫基磺酸甲酯（MMTS）用于实验生物化学中，以使蛋白半胱氨酸的巯基烷基化。它的应用主要包括捕获氧化还原敏感蛋白和经过S-亚硝基化的蛋白的天然硫醇-二硫键状态。该试剂也可以用作酶活性的抑制剂，因为亲核半胱氨酸硫醇盐通常存在于各种酶的活性位点。与使用常规试剂的不可逆烷基化相比，为此目的使用MMTS的优点是可形成甲硫基混合二硫化物的可逆性。其他优点包括MMTS的尺寸小以及保护和脱保护程序的简单性，使其可以很好地接近掩埋的蛋白半胱氨酸。在这项研究中，我们报告了MMTS在涉及氧化还原酶（甘油醛-3-磷酸脱氢酶，GAPDH），氧化还原调节蛋白（recoverin）和半胱氨酸蛋白酶（triticain-α）的实验中的应用实例。我们证明，一方面，MMTS可以修饰巯基酶GAPDH中的功能性半胱氨酸，从而防止巯基氧化并可逆地抑制该酶，另一方面，它可以保护Recoveryin的氧化还原敏感巯基免受氧化，这种修饰产生对蛋白质活性没有影响。此外，以木瓜蛋白酶样酶Triticain-α为例，我们报道了MMTS在长期纯化和复性过程中作为活性半胱氨酸蛋白酶一级结构的保护剂的新应用。根据数据，