Poljak Anne, Grant Ross, Austin Chris J D, Jamie Joanne F, Willows Robert D, Takikawa Osamu, Littlejohn Tamantha K, Truscott Roger J W, Walker Mark J, Sachdev Perminder, Smythe George A
Bioanalytical Mass Spectrometry Facility, University of New South Wales, Sydney, Australia.
Arch Biochem Biophys. 2006 Jun 1;450(1):9-19. doi: 10.1016/j.abb.2006.03.003. Epub 2006 Mar 20.
Indoleamine 2,3-dioxygenase is the first and rate limiting enzyme of the kynurenine pathway of tryptophan metabolism, has potent effects on cell proliferation and mediates antimicrobial, antitumorogenic, and immunosuppressive effects. As a potent cytotoxic effector, the mechanisms of indoleamine 2,3-dioxygenase inhibition deserve greater attention. The work presented here represents the first systematic study exploring the mechanisms by which low levels of hydrogen peroxide (10-100 microM) inhibit indoleamine 2,3-dioxygenase in vitro. Following brief peroxide exposure both enzyme inhibition and structural changes were observed. Loss of catalysis was accompanied by oxidation of several cysteine residues to sulfinic and sulfonic acids, observed by electrospray and MALDI mass spectrometry. Enzyme activity could in part be preserved in the presence of sulfhydryl containing compounds, particularly DTT and methionine. However, these structural alterations did not prevent substrate (l-tryptophan) binding. Some enzyme activity could be recovered in the presence of thioredoxin, indicating that the inhibitory effect of H(2)O(2) is at least partially reversible in vitro. We present evidence that cysteine oxidation represents one mechanism of indoleamine 2,3-dioxygenase inhibition.
吲哚胺2,3-双加氧酶是色氨酸代谢犬尿氨酸途径的首个限速酶，对细胞增殖具有显著作用，并介导抗菌、抗肿瘤和免疫抑制作用。作为一种有效的细胞毒性效应物，吲哚胺2,3-双加氧酶抑制机制值得更多关注。本文介绍的工作是首次系统研究低水平过氧化氢（10 - 100微摩尔）在体外抑制吲哚胺2,3-双加氧酶的机制。短暂暴露于过氧化物后，观察到酶抑制和结构变化。通过电喷雾和基质辅助激光解吸电离质谱法观察到，催化作用丧失伴随着几个半胱氨酸残基氧化为亚磺酸和磺酸。在含有巯基的化合物，特别是二硫苏糖醇和蛋氨酸存在的情况下，酶活性部分得以保留。然而，这些结构改变并未阻止底物（L-色氨酸）结合。在硫氧还蛋白存在的情况下，部分酶活性可以恢复，这表明过氧化氢的抑制作用在体外至少部分是可逆的。我们提供的证据表明，半胱氨酸氧化是吲哚胺2,3-双加氧酶抑制的一种机制。
