Detection of a C4a-Hydroperoxyflavin Intermediate in the Reaction of a Flavoprotein Oxidase

• Published in: Biochemistry (Easton) Vol. 47; no. 33; pp. 8485 - 8490
• Main Authors: Sucharitakul, Jeerus, Prongjit, Methinee, Haltrich, Dietmar, Chaiyen, Pimchai
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 19-08-2008
• Subjects: Basidiomycota - enzymology; Flavins - chemistry; Flavins - metabolism; Flavoproteins - metabolism; Kinetics; Models, Molecular; Molecular Structure; Oxidation-Reduction; Oxidoreductases - chemistry; Oxidoreductases - metabolism
• ISSN: 0006-2960; 1520-4995
• DOI: 10.1021/bi801039d

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 × 104 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.