Maeda-Yorita K, Aki K, Sagai H, Misaki H, Massey V
Institute for Enzyme Research, University of Tokushima, Japan.
Biochimie. 1995;77(7-8):631-42. doi: 10.1016/0300-9084(96)88178-8.
Properties of L-lactate oxidase from Aerococcus viridans are described. The gene encoding the enzyme has been isolated. From its cDNA sequence the amino acid sequence has been derived and shown to have high similarity with those of other enzymes catalyzing oxidation of L-alpha-hydroxy acids, including flavocytochrome b2, lactate monooxygenase, glycolate oxidase, mandelate dehydrogenases and a long chain alpha-hydroxy acid oxidase. The enzyme is expressed in Escherichia coli, and is a flavoprotein containing FMN as prosthetic group. It shares many properties of other alpha-hydroxy acid oxidizing enzymes, eg stabilization of the anionic semiquinone form of the flavin, facile formation of flavin-N(5)-sulfite adducts and a set of conserved amino acid residues around the bound flavin. Steady-state and rapid reaction kinetics of the enzyme have been studied and found to share many characteristics with those of L-lactate monooxygenase, but to differ from the latter in quantitative aspects. It is these quantitative differences between the two enzymes which account for the differences in the overall reactions catalyzed. These differences arise from different stabilities of a common intermediate of reduced flavin enzyme and pyruvate. In the case of the monooxygenase this complex is very stable and is the form that reacts with O2 to give a complex in which the oxidative decarboxylation occurs, yielding the products, acetate, CO2, and H2O (Lockridge O, Massey V, Sullivan PA (1972) J Biol Chem 247, 8097-8106). With lactate oxidase, the complex dissociates rapidly, with the result that it is the free reduced flavin form of the enzyme that reacts with O2, to give the observed products, pyruvate and H2O2.
本文描述了绿色气球菌中L-乳酸氧化酶的特性。编码该酶的基因已被分离。从其cDNA序列推导得到了氨基酸序列，结果表明该序列与其他催化L-α-羟基酸氧化的酶具有高度相似性，这些酶包括黄素细胞色素b2、乳酸单加氧酶、乙醇酸氧化酶、扁桃酸脱氢酶和长链α-羟基酸氧化酶。该酶在大肠杆菌中表达，是一种以FMN为辅基的黄素蛋白。它与其他α-羟基酸氧化酶具有许多共同特性，例如黄素阴离子半醌形式的稳定化、黄素-N(5)-亚硫酸盐加合物的容易形成以及结合黄素周围的一组保守氨基酸残基。研究了该酶的稳态和快速反应动力学，发现其与L-乳酸单加氧酶具有许多共同特征，但在定量方面有所不同。正是这两种酶之间的定量差异导致了它们所催化的整体反应的差异。这些差异源于还原黄素酶和丙酮酸的共同中间体的不同稳定性。对于单加氧酶来说，这种复合物非常稳定，并且是以这种形式与O2反应生成一种复合物，在该复合物中发生氧化脱羧反应，产生产物乙酸盐、CO2和H2O（Lockridge O, Massey V, Sullivan PA (1972) J Biol Chem 247, 8097 - 8106）。而对于乳酸氧化酶，该复合物迅速解离，结果是酶的游离还原黄素形式与O2反应，生成观察到的产物丙酮酸和H2O2。
