Quinone-induced protein modifications: Kinetic preference for reaction of 1,2-benzoquinones with thiol groups in proteins
Quinone-induced protein modifications: Kinetic preference for reaction of 1,2-benzoquinones with thiol groups in proteins
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醌诱导的蛋白质修饰：1,2-苯醌与蛋白质中硫醇基团反应的动力学偏好
DOI：
10.1016/j.freeradbiomed.2016.05.019
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发表时间：
2016-08-01
影响因子：
7.4
通讯作者：
Lund, Marianne N.
中科院分区：
文献类型：
--
作者：
Li, Yuting;Jongberg, Sisse;Lund, Marianne N.
多酚氧化成醌是一种抗氧化机制，但所产生的醌可能会对蛋白质造成损伤，因为它们通过迈克尔加成与亲核基团（如硫醇和胺）反应生成蛋白质加合物。采用紫外-可见停流光度法测定了4-甲基苯醌（4 MBQ）与蛋白质、硫醇和胺类化合物在准一级反应条件下的反应速率常数。通过LC-ESI-MS/MS鉴定了加合物的化学结构。具有游离巯基的蛋白质被4 MBQ快速修饰，在pH 7.0下，牛血清白蛋白（BSA）和人血清白蛋白的表观二级速率常数k（2）分别为（3.1 +/- 0.2）x 10（4）M-1 s（-1）和（4.8 +/- 0.2）x 10（2）M-1 s（-1）。这些值至少是α-乳清蛋白（4.0 +/- 0.2）x 10（2）M-1 s（-1）的12倍，后者不含任何游离巯基。BSA的Cys-34与N-乙基马来酰亚胺的反应将硫醇浓度降低了约59%，这导致k（2）降低了约59%，这与Cys-34处的快速加合一致。4 MBQ与胺（Gly、N α-乙酰基-L-Lys、N ε-乙酰基-L-Lys和L-Lys）和N α-乙酰基-L-Arg的胍基的反应比与低分子量硫醇（L-Cys、N α-乙酰基-L-Cys、谷胱甘肽）的反应慢至少5 × 10（5）。硫醇-醌相互作用通过中间加合物形成无色的硫醇-苯酚产物，而胺-醌相互作用产生需要氧参与的有色胺-醌产物。这些数据提供了强有力的证据，快速修改蛋白质巯基醌物种，这可能是相当重要的生物和食品系统。(C)2016 Elsevier Inc. All rights reserved.
Oxidation of polyphenols to quinones serves as an antioxidative mechanism, but the resulting quinones may induce damage to proteins as they react through a Michael addition with nucleophilic groups, such as thiols and amines to give protein adducts. In this study, rate constants for the reaction of 4-methylbenzoquinone (4MBQ) with proteins, thiol and amine compounds were determined under pseudo first order conditions by UV-vis stopped-flow spectrophotometry. The chemical structures of the adducts were identified by LC-ESI-MS/MS.Proteins with free thiols were rapidly modified by 4MBQ with apparent second order rate constants, k(2) of (3.1 +/- 0.2) x 10(4) M-1 s(-1) for bovine serum albumin (BSA) and (4.8 +/- 0.2) x 10(2) M-1 s(-1) for human serum albumin at pH 7.0. These values are at least 12-fold greater than that for alpha-lactalbumin (4.0 +/- 0.2) x 10(2) M-1 s(-1), which does not contain any free thiols. Reaction of Cys-34 of BSA with N-ethylmaleimide reduced the thiol concentration by similar to 59%, which resulted in a decrease in k(2) by a similar percentage, consistent with rapid adduction at Cys-34. Reaction of 4MBQ with amines (Gly, N alpha-acetyl-L-Lys, N epsilon-acetyl-L-Lys and L-Lys) and the guanidine group of N alpha-acetyl-L-Arg was at least 5 x 10(5) slower than with low-molecular-mass thiols (L-Cys, N alpha-acetyl-L-Cys, glutathione). The thiol-quinone interactions formed colorless thiol-phenol products via an intermediate adduct, while the amine-quinone interactions generated colored amine-quinone products that require oxygen involvement. These data provide strong evidence for rapid modification of protein thiols by quinone species which may be of considerable significance for biological and food systems. (C) 2016 Elsevier Inc. All rights reserved.