Detection of a C4a-hydroperoxyflavin intermediate in the reaction of a flavoprotein oxidase.
This work describes for the first time the identification of a reaction intermediate, C4a-hydroperoxyflavin, during the oxidative half-reaction of a flavoprotein oxidase, pyranose 2-oxidase (P2O) from Trametes multicolor, by using rapid kinetics. The reduced P2O reacted with oxygen with a forward rate constant of 5.8 x 10 (4) M (-1) s (-1) and a reverse rate constant of 2 s (-1), resulting in the formation of a C4a-hydroperoxyflavin intermediate which decayed with a rate constant of 18 s (-1). The absorption spectrum of the intermediate resembled the spectra of flavin-dependent monooxygenases. A hydrophobic cavity formed at the re side of the flavin ring in the closed state structure of P2O may help in stabilizing the intermediate.
Sucharitakul
Jeerus
J
Department of Biochemistry, Faculty of Dentistry, Chulalongkorn University, Henri-Dunant Road, Patumwan, Bangkok 10300, Thailand.
Prongjit
Methinee
M
Haltrich
Dietmar
D
Chaiyen
Pimchai
P
D001487:Basidiomycota / Q000201:enzymology; D005415:Flavins / Q000737:chemistry* / Q000378:metabolism*; D005420:Flavoproteins / Q000378:metabolism*; D007700:Kinetics; D008958:Models, Molecular; D015394:Molecular Structure; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000737:chemistry / Q000378:metabolism*
C531259:C4a-hydroperoxyflavin; D005415:Flavins; D005420:Flavoproteins; D010088:Oxidoreductases