Potentiometric analysis of UDP-galactopyranose mutase: Stabilization of the flavosemiquinone by substrate
Potentiometric analysis of UDP-galactopyranose mutase: Stabilization of the flavosemiquinone by substrate
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DOI：
10.1021/bi027077f
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发表时间：
2003-02-25
期刊：
影响因子：
2.9
通讯作者：
Naismith, JH
中科院分区：
文献类型：
--
作者：
Fullerton, SWB;Daff, S;Naismith, JH
UDP-吡喃半乳糖变位酶是一种黄素蛋白，其催化UDP-吡喃半乳糖和UDP-呋喃半乳糖的相互转化。这种酶之所以令人感兴趣，是因为它提供了呋喃半乳糖的活化生物合成前体，呋喃半乳糖是许多细菌病原体的关键细胞壁成分。这种变位酶的反应机制很有趣，因为异头氧形成糖苷键，这意味着反应必须通过涉及环断裂和闭合的新机制进行。该酶的结构是已知的，但其机制虽然有所推测，但尚未解决。总体反应是电中性的，但黄素必须采用还原形式才能发挥活性，这表明隐氧化还原反应。在此，我们报告了酶黄素辅因子的热力学分析，目的是定义系统并设置可能的反应方案的参数。分析表明，中性半醌(FADH(.))在底物存在下稳定，完全还原的黄素是阴离子FADH(-)而不是中性FADH(2)。阴离子 FADH(-) 有潜力充当快速 1 电子供体/受体，而不会因耦合质子转移而减慢，因此是理想的隐氧化还原辅助因子。
UDP-galactopyranose mutase is a flavoprotein which catalyses the interconversion of UDP-galactopyranose and UDP-galactofuranose. The enzyme is of interest because it provides the activated biosynthetic precursor of galactofuranose, a key cell wall component of many bacterial pathogens. The reaction mechanism of this mutase is intriguing because the anomeric oxygen forms a glycosidic bond, which means that the reaction must proceed by a novel mechanism involving ring breakage and closure. The structure of the enzyme is known, but the mechanism, although speculated on, is not resolved. The overall reaction is electrically neutral but a crypto-redox reaction is suggested by the requirement that the flavin must adopt the reduced form for activity. Herein we report a thermodynamic analysis of the enzyme's flavin cofactor with the objective of defining the system and setting parameters for possible reaction schemes. The analysis shows that the neutral semiquinone (FADH(.)) is stabilized in the presence of substrate and the fully reduced flavin is the anionic FADH(-) rather than the neutral FADH(2). The anionic FADH(-) has the potential to act as a rapid 1-electron donor/acceptor without being slowed by a coupled proton transfer and is therefore an ideal crypto-redox cofactor.