Ebselen modulates target proteins through redox reactions with selenocysteine/cysteine residues, or through binding to the zinc finger domains. However, a recent contradiction in ebselen inhibition of kidney type glutaminase (KGA) stimulated our interest in investigating its inhibition mechanism with glutamate dehydrogenase (GDH), KGA, thioredoxin reductase (TrxR), and glutathione S-transferase. Fluorescein- or biotin-labeled ebselen derivatives were synthesized for mechanistic analyses. Biomolecular interaction analyses showed that only GDH, KGA, and TrxR proteins can bind to the ebselen derivative, and the binding to GDH and KGA could be competed off by glutamine or glutamate. From the gel shift assays, the fluorescein-labeled ebselen derivative could co-migrate with hexameric GDH and monomeric/dimeric TrxR in a dose-dependent manner; it also co-migrated with KGA but disrupted the tetrameric form of the KGA enzyme at a high compound concentration. Further proteomic analysis demonstrated that the ebselen derivative could cross-link with proteins through a specific cysteine at the active site of GDH and TrxR proteins, but for KGA protein, the binding site is at the N-terminal appendix domain outside of the catalytic domain, which might explain why ebselen is not a potent KGA enzyme inhibitor in functional assays. In conclusion, ebselen could inhibit enzyme activity by binding to the catalytic domain or disruption of the protein complex. In addition, ebselen is a relatively potent selective GDH inhibitor that might provide potential therapeutic opportunities for hyperinsulinism-hyperammonemia syndrome patients who have the mutational loss of GTP inhibition.

中文翻译：

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Ebselen：谷氨酸脱氢酶和谷氨酰胺酶抑制的机制

Ebselen通过与硒代半胱氨酸/半胱氨酸残基的氧化还原反应，或通过与锌指结构域的结合来调节靶蛋白。但是，最近在对ebselen抑制肾型谷氨酰胺酶（KGA）的矛盾中激发了我们研究谷氨酸脱氢酶（GDH），KGA，硫氧还蛋白还原酶（TrxR）和谷胱甘肽S的抑制机制的兴趣。-转移酶。合成了荧光素或生物素标记的依布硒啉衍生物以进行机械分析。生物分子相互作用分析表明，只有GDH，KGA和TrxR蛋白可以与依ebselen衍生物结合，而与GDH和KGA的结合可以被谷氨酰胺或谷氨酸竞争。从凝胶位移分析中，荧光素标记的依布硒啉衍生物可以与六聚体GDH和单体/二聚体TrxR共同迁移，且呈剂量依赖性。它也与KGA共同迁移，但在高化合物浓度下破坏了KGA酶的四聚体形式。进一步的蛋白质组学分析表明，依布硒啉衍生物可通过GDH和TrxR蛋白质活性位点上的特定半胱氨酸与蛋白质交联，但对于KGA蛋白质，结合位点位于催化结构域之外的N末端阑尾结构域，这可能可以解释为什么依布硒仑在功能测定中不是有效的KGA酶抑制剂。总之，依布硒仑可以通过与催化结构域结合或破坏蛋白质复合物来抑制酶的活性。此外，依布硒仑是一种相对有效的选择性GDH抑制剂，可能为具有GTP抑制突变突变的高胰岛素血症-高氨血症综合征患者提供潜在的治疗机会。