L-lactate oxidase and L-lactate monooxygenase: mechanistic variations on a common structural theme.
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.
Maeda-Yorita
K
K
Institute for Enzyme Research, University of Tokushima, Japan.
Aki
K
K
Sagai
H
H
Misaki
H
H
Massey
V
V
D000595:Amino Acid Sequence; D002384:Catalysis; D004795:Enzyme Stability; D006706:Homeostasis; D007700:Kinetics; D006899:Mixed Function Oxygenases / Q000737:chemistry*; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D010100:Oxygen / Q000737:chemistry; D011773:Pyruvates / Q000737:chemistry; D019289:Pyruvic Acid; D017386:Sequence Homology, Amino Acid; D013053:Spectrophotometry; D013056:Spectrophotometry, Ultraviolet; D013447:Sulfites / Q000737:chemistry
D011773:Pyruvates; D013447:Sulfites; D019289:Pyruvic Acid; D006899:Mixed Function Oxygenases; C021497:lactate 2-monooxygenase; D010100:Oxygen