Ariza Antonio, Vickers Tim J, Greig Neil, Armour Kirsten A, Dixon Mark J, Eggleston Ian M, Fairlamb Alan H, Bond Charles S
Division of Biological Chemistry & Molecular Microbiology, Wellcome Trust Biocentre, School of Life Sciences, University of Dundee, Dundee DD1 5EH, Scotland, UK.
Mol Microbiol. 2006 Feb;59(4):1239-48. doi: 10.1111/j.1365-2958.2006.05022.x.
Trypanothione replaces glutathione in defence against cellular damage caused by oxidants, xenobiotics and methylglyoxal in the trypanosomatid parasites, which cause trypanosomiasis and leishmaniasis. In Leishmania major, the first step in methylglyoxal detoxification is performed by a trypanothione-dependent glyoxalase I (GLO1) containing a nickel cofactor; all other characterized eukaryotic glyoxalases use zinc. In kinetic studies L. major and human enzymes were active with methylglyoxal derivatives of several thiols, but showed opposite substrate selectivities: N1-glutathionylspermidine hemithioacetal is 40-fold better with L. major GLO1, whereas glutathione hemithioacetal is 300-fold better with human GLO1. Similarly, S-4-bromobenzylglutathionylspermidine is a 24-fold more potent linear competitive inhibitor of L. major than human GLO1 (Kis of 0.54 microM and 12.6 microM, respectively), whereas S-4-bromobenzylglutathione is >4000-fold more active against human than L. major GLO1 (Kis of 0.13 microM and >500 microM respectively). The crystal structure of L. major GLO1 reveals differences in active site architecture to both human GLO1 and the nickel-dependent Escherichia coli GLO1, including increased negative charge and hydrophobic character and truncation of a loop that may regulate catalysis in the human enzyme. These differences correlate with the differential binding of glutathione and trypanothione-based substrates, and thus offer a route to the rational design of L. major-specific GLO1 inhibitors.
在引起锥虫病和利什曼病的锥虫目寄生虫中，三巯基丙胺（trypanothione）替代了谷胱甘肽（glutathione），以抵御由氧化剂、异生物素和甲基乙二醛引起的细胞损伤。在硕大利什曼原虫（Leishmania major）中，甲基乙二醛解毒的第一步由含有镍辅因子的三巯基丙胺依赖性乙二醛酶I（GLO1）完成；所有其他已表征的真核生物乙二醛酶都使用锌。在动力学研究中，硕大利什曼原虫和人类的酶对几种硫醇的甲基乙二醛衍生物都有活性，但表现出相反的底物选择性：N1-谷胱甘肽亚精胺半硫代乙缩醛对硕大利什曼原虫GLO1的活性高40倍，而谷胱甘肽半硫代乙缩醛对人类GLO1的活性高300倍。同样，S-4-溴苄基谷胱甘肽亚精胺对硕大利什曼原虫的线性竞争性抑制作用比对人类GLO1强24倍（Kis分别为0.54 microM和12.6 microM），而S-4-溴苄基谷胱甘肽对人类的活性比对硕大利什曼原虫GLO1高>4000倍（Kis分别为0.13 microM和>500 microM）。硕大利什曼原虫GLO1的晶体结构揭示了其与人类GLO1和镍依赖性大肠杆菌GLO1在活性位点结构上的差异，包括负电荷和疏水性增加以及一个可能调节人类酶催化作用的环的截断。这些差异与基于谷胱甘肽和三巯基丙胺的底物的差异结合相关，因此为合理设计硕大利什曼原虫特异性GLO1抑制剂提供了一条途径。
