Peptidylglycine-alpha-hydroxylating monooxygenase generates two hydroxylated products from its mechanism-based suicide substrate, 4-phenyl-3-butenoic acid.
Peptidylglycine-alpha-hydroxylating monooxygenase generates two hydroxylated products from its mechanism-based suicide substrate, 4-phenyl-3-butenoic acid.
复制标题
肽基甘氨酸-α-羟基化单加氧酶从其基于机制的自杀底物 4-苯基-3-丁烯酸产生两种羟基化产物。
DOI：
10.1021/bi0002380
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发表时间：
2000
期刊：
影响因子：
2.9
通讯作者：
Mueller,GP
中科院分区：
文献类型：
--
作者：
Driscoll,WJ;König,S;Fales,HM;Pannell,LK;Eipper,BA;Mueller,GP
双功能酶肽基甘氨酸-α-酰胺化单加氧酶介导 C 末端甘氨酸延伸肽转化为其活性 α-酰胺化产物。肽基甘氨酸-α-羟基化单加氧酶（PHM，EC 1.14.17.3）催化该两步过程中的第一个反应。烯属化合物 4-苯基-3-丁烯酸 (PBA) 是已知最有效的不可逆、基于机制的 PHM 灭活剂。虽然 PBA 对 PHM 抑制作用的细节仍不清楚，但已提出蛋白质的共价修饰作为潜在机制。我们在此报道，在灭活 PHM 的过程中，PBA 本身充当底物，而不共价标记酶。在饱和条件下，每个失活的 PHM 分子大约代谢 100 个 PBA 分子。 PBA 通过 PHM 代谢产生两种羟基化产物，2-羟基-4-苯基-3-丁烯酸及其烯丙基异构体，4-羟基-4-苯基-2-丁烯酸。虽然每种产物的一种对映体在反应中明显受到青睐，但两种对映体都会产生。从这些观察中，我们得出结论，羟基化 PBA 产物是通过离域自由基机制形成的，并且缺乏绝对立体特异性表明催化位点内具有显着的运动自由度。 PHM 代谢 PBA 的能力表明 PHM 的生理功能可能包括除含有末端甘氨酸的底物之外的底物的羟基化。
The bifunctional enzyme peptidylglycine-α-amidating monooxygenase mediates the conversion of C-terminal glycine-extended peptides to their active α-amidated products. Peptidylglycine-α-hydroxylating monooxygenase (PHM, EC 1.14.17.3) catalyzes the first reaction in this two-step process. The olefinic compound 4-phenyl-3-butenoic acid (PBA) is the most potent irreversible, mechanism-based PHM inactivator known. While the details of the inhibitory action of PBA on PHM remain undefined, covalent modification of the protein has been proposed as the underlying mechanism. We report here that, in the process of inactivating PHM, PBA itself serves as a substrate without covalently labeling the enzyme. Approximately 100 molecules of PBA are metabolized per molecule of PHM inactivated, under saturating conditions. The metabolism of PBA by PHM generates two hydroxylated products, 2-hydroxy-4-phenyl-3-butenoic acid and its allylic isomer, 4-hydroxy-4-phenyl-2-butenoic acid. While one enantiomer for each product is significantly favored in the reaction, both are produced. From these observations, we conclude that hydroxylated PBA products are formed by a delocalized free radical mechanism and that the lack of absolute stereospecificity indicates significant freedom of movement within the catalytic site. The ability of PHM to metabolize PBA suggests that the physiological functions of PHM may include the hydroxylation of substrates other than those containing terminal glycines.