Barshteyn Nella, Elfarra Adnan A
Department of Comparative Biosciences, University of Wisconsin, Madison 53706, USA.
Chem Res Toxicol. 2007 Oct;20(10):1563-9. doi: 10.1021/tx700263w. Epub 2007 Sep 25.
Previously, our laboratory has shown that S-(1,2-dichlorovinyl)-L-cysteine sulfoxide (DCVCS), a Michael acceptor produced by a flavin-containing monooxygenase 3 (FMO3)-mediated oxidation of S-(1,2-dichlorovinyl)-L-cysteine (DCVC), is a more potent nephrotoxicant than DCVC. In the present study, we characterized reactions of DCVCS with nucleophilic amino acids. DCVCS incubations with N-acetyl-L-cysteine (NAC) at pH 7.4 and 37 degrees C for 1 h resulted in the formation of three monoadducts and one diadduct characterized by LC/MS, 1H NMR, and 1H-detected heteronuclear single quantum correlation. The formation of all adducts (with relative ratios of 29, 31, 24, and 12%, respectively) was rapid and time-dependent; the half-lives of the two DCVCS diastereomers in the presence of NAC were 13.8 (diastereomer I) and 9.4 min (diastereomer II). Adducts 1 and 2 were determined to be diastereomers of S-[1-chloro-2-(N-acetyl-L-cystein- S-yl)vinyl]-L-cysteine sulfoxide formed by Michael addition of NAC to the terminal vinylic carbon of DCVCS followed by loss of HCl. Adduct 4 was determined to be S-[2-chloro-2-(N-acetyl-L-cystein- S-yl)vinyl]-L-cysteine sulfoxide formed from the initial Michael addition product followed by a less favorable loss of HCl and/or by a rearrangement of adduct 2 through the formation of a cyclic chloronium ion. The addition of another molecule of NAC to monoadducts 1, 2, or 4 resulted in the formation of the novel diadduct, S-[2,2-( N-acetyl-L-cystein-S-yl)vinyl]-L-cysteine sulfoxide (adduct 3), whose detection in relatively large amount suggests that DCVCS could act as a cross-linking agent. DCVCS was not reactive with N-acetyl-L-lysine or L-valinamide at similar incubation conditions. Collectively, the results suggest selective reactivity of DCVCS toward protein sulfhydryl groups. Furthermore, the cross-linking properties of DCVCS may in part explain its high nephrotoxic potency.
此前，我们实验室已表明，S-(1,2-二氯乙烯基)-L-半胱氨酸亚砜（DCVCS）是一种由含黄素单加氧酶3（FMO3）介导的S-(1,2-二氯乙烯基)-L-半胱氨酸（DCVC）氧化产生的迈克尔受体，它是一种比DCVC更强效的肾毒物。在本研究中，我们对DCVCS与亲核氨基酸的反应进行了表征。在pH 7.4和37℃条件下，将DCVCS与N-乙酰-L-半胱氨酸（NAC）孵育1小时，通过液相色谱/质谱、1H核磁共振和1H检测的异核单量子相关表征，结果形成了三种单加合物和一种双加合物。所有加合物的形成（相对比例分别为29%、31%、24%和12%）都很快且具有时间依赖性；在NAC存在下，两种DCVCS非对映异构体的半衰期分别为13.8分钟（非对映异构体I）和9.4分钟（非对映异构体II）。加合物1和2被确定为S-[1-氯-2-(N-乙酰-L-半胱氨酰-S-基)乙烯基]-L-半胱氨酸亚砜的非对映异构体，它是由NAC对DCVCS末端乙烯基碳进行迈克尔加成，随后失去HCl形成的。加合物4被确定为S-[2-氯-2-(N-乙酰-L-半胱氨酰-S-基)乙烯基]-L-半胱氨酸亚砜，它由最初的迈克尔加成产物形成，随后不太有利地失去HCl和/或通过形成环状氯鎓离子使加合物2发生重排。向单加合物1、2或4中再加入一分子NAC会导致形成新型双加合物S-[2,2-(N-乙酰-L-半胱氨酰-S-基)乙烯基]-L-半胱氨酸亚砜（加合物3），其相对大量的检测表明DCVCS可能作为一种交联剂。在类似的孵育条件下，DCVCS与N-乙酰-L-赖氨酸或L-缬氨酰胺无反应。总体而言，结果表明DCVCS对蛋白质巯基具有选择性反应性。此外，DCVCS的交联特性可能部分解释了其高肾毒性效力。
