Methylarsenicals and arsinothiols are potent inhibitors of mouse liver thioredoxin reductase
Methylarsenicals and arsinothiols are potent inhibitors of mouse liver thioredoxin reductase
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DOI：
10.1021/tx9900775
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发表时间：
1999-10-01
影响因子：
4.1
通讯作者：
Thomas, DJ
中科院分区：
文献类型：
--
作者：
Lin, S;Cullen, WR;Thomas, DJ
硫氧还蛋白还原酶（TR，EC 1.6.4.5）从成年雄性B6 C3 F1小鼠的肝脏纯化5800倍。纯化的蛋白质的估计分子量为约57 kDa。纯化的酶的活性通过5，5 '-二硫代双（2-硝基苯甲酸）（DTNB）的NADPH依赖性还原来监测;该活性被1 μ M金硫葡萄糖完全抑制。砷和arsinothiols，作为-III-含化合物与L-半胱氨酸或谷胱甘肽的复合物，进行了测试作为纯化的酶的DTNB还原酶活性的抑制剂。五价砷的抑制作用远低于三价砷。在所有砷剂中，CH_3As Ⅲ是TR最强的抑制剂。发现CH 3AsIII是DTNB还原的竞争性抑制剂（Ki类似于100 nM）和NADPH氧化的非竞争性抑制剂。TR的抑制CH 3AsIII是时间依赖性的，不能通过添加一个含二硫醇的分子，2，3-二巯基丁二酸，反应混合物中逆转。TR的抑制CH 3AsIII需要在反应混合物中的NADPH的同时存在。然而，与其他吡啶核苷酸二硫键氧化还原酶不同，没有证据表明小鼠肝脏TR因暴露于NADPH而失活。用CH 3As III处理没有增加纯化的酶的NADPH氧化酶活性。因此，CH 3AsIII，一个假定的中间体的生物甲基化的途径，是一种有效的和不可逆的抑制剂的酶参与的细胞对氧化应激的反应。
Thioredoxin reductase (TR, EC 1.6.4.5) was purified 5800-fold from the livers of adult male B6C3F1 mice. The estimated molecular mass of the purified protein was about 57 kDa, The activity of the purified enzyme was monitored by the NADPH-dependent reduction of 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB); this activity was fully inhibited by 1 mu M aurothioglucose. Arsenicals and arsinothiols, complexes of As-III-containing compounds with L-cysteine or glutathione, were tested as inhibitors of the DTNB reductase activity of the purified enzyme. Pentavalent arsenicals were much less potent inhibitors than trivalent arsenicals. Among all the arsenicals, CH3AsIII was the most potent inhibitor of TR. CH3AsIII was found to be a competitive inhibitor of the reduction of DTNB (K-i similar to 100 nM) and a noncompetitive inhibitor of the oxidation of NADPH. The inhibition of TR by CH3AsIII was time-dependent and could not be reversed by the addition of a dithiol-containing molecule, 2,3-dimercaptosuccinic acid, to the reaction mixture. The inhibition of TR by CH3AsIII required the simultaneous presence of NADPH in the reaction mixture. However, unlike other pyridine nucleotide disulfide oxidoreductases, there was no evidence that mouse liver TR was inactivated by exposure to NADPH. Treatment with CH3AsIII did not increase the NADPH oxidase activity of the purified enzyme. Thus, CH3AsIII, a putative intermediate in the pathway for the biomethylation of As, is a potent and irreversible inhibitor of an enzyme involved in the response of the cell to oxidative stress.