Naslov
Reaction of Trigonopsis variabilis D-amino acid oxidase with 2, 6-dichloroindophenol: kinetic characterization and development of an oxygen-independent assay of the enzyme activity

Autori
Trampitsch, Christian ; Slavica, Anita ; Riethorst, Waander ; Nidetzky, Bernd

Izvornik
Journal of Molecular Catalysis B - Enzymatic (1381-1177) 32 (2005), 5-6; 271-278

Vrsta, podvrsta i kategorija rada
Radovi u časopisima, članak, znanstveni

Ključne riječi
flavoenzyme; electron acceptor; oxygen; biocatalysis; enzyme assay

Sažetak
2, 6-Dichloroindophenol (DCIP) is shown to be utilised efficientlz as electron acceptor replacing dioxygen in the reaction of Trigonopsis variabilis D-amino acid oxidase (TvDAO) with D-Methionine as the susbstrate. The specifificty constant for DCIP reduction at 30 C is one twelfth that of oxygen conversion into hydrogen peroxide. Time course analysis of symultaneous consumption of DCIP and dioxygen, recorded on-line by absorption and non-invasive fluorescence quenching, respectively, pinpoints the preferential utilisation of dioxygen ; and reveals a maximum DCIP conversion rate that is independent of the initial concentration of dioxygen. A robust direct assay of TvDAO activity has been developed that does not require ananerobic reaction conditions. It was down-scaled to microtitre plate format and overcomes practical limitations of other assays due to low affnity of TvDAO for dioxygen (Km  0.7 mmol L-1).

Izvorni jezik
Engleski

Znanstvena područja
Biotehnologija

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