Hiltunen J K, Palosaari P M, Kunau W H
Abteilung für Zellbiochemie, Medizinische Fakultät, Ruhr-Universität Bochum, Federal Republic of Germany.
J Biol Chem. 1989 Aug 15;264(23):13536-40.
Interconversion of D- and L-isomers of 3-hydroxy-decanoyl-CoA was catalyzed by rat liver homogenate. Cation exchange chromatography followed by ammonium sulfate precipitation and PBE-94 chromatofocusing column was used to separate the peroxisomal bifunctional protein, the classic 2-enoyl-CoA hydratase (crotonase), and a novel 2-enoyl-CoA hydratase. Epimerization activity was lost during the last purification step. None of the above proteins was capable of catalyzing the epimerization by itself, but reconstitution was achieved by recombining crotonase and the novel 2-enoyl-CoA hydratase. Since hydration by the latter enzyme follows a different stereochemical course from that with crotonase, these two hydratases are distinguished as 2-enoyl-CoA hydratase 1 (crotonase) and 2-enoyl-CoA hydratase 2 (the novel hydratase). The data strongly suggested that epimerization in the rat liver proceeds via dehydration-hydration catalyzed by the two different hydratases. The intermediate of this two step mechanism appears to be trans-2-enoyl-CoA.
大鼠肝脏匀浆可催化3-羟基癸酰辅酶A的D型和L型异构体的相互转化。采用阳离子交换色谱法，随后进行硫酸铵沉淀和PBE-94色谱聚焦柱分离过氧化物酶体双功能蛋白、经典的2-烯酰辅酶A水合酶（巴豆酸酶）和一种新型的2-烯酰辅酶A水合酶。在最后一步纯化过程中，差向异构化活性丧失。上述蛋白质自身均无催化差向异构化的能力，但通过重组巴豆酸酶和新型2-烯酰辅酶A水合酶可实现重组。由于后一种酶的水合作用遵循与巴豆酸酶不同的立体化学过程，这两种水合酶分别被区分为2-烯酰辅酶A水合酶1（巴豆酸酶）和2-烯酰辅酶A水合酶2（新型水合酶）。数据强烈表明，大鼠肝脏中的差向异构化是通过两种不同水合酶催化的脱水-水合作用进行的。这一两步机制的中间产物似乎是反式-2-烯酰辅酶A。
