A steady-state kinetic study of the reaction catalysed by the secondary-amine mono-oxygenase of Pseudomonas aminovorans.
1. Secondary-amine mono-oxygenase (proposed EC group 1.14.99.-) was partially purified from trimethylamine-grown Pseudomonas aminovorans by (NH4)2SO4 fractionation, gel filtration, hydrophobic chromatography on 5-aminopentylamino-Sepharose, and affinity chromatography on Sepharose-bound NADH. 2. Some problems in the affinity-chromatography step are discussed. 3. A steady-state kinetic analysis varying substrate, oxygen and electron-donor concentrations was performed, which, over the concentration range studied, gave a series of families of approximately parallel double-reciprocal plots. From secondary and tertiary plots, Michaelis constants of 0.160 mM, 0.086 mM and 0.121 mM were obtained for dimethylamine, NADPH and oxygen respectively. 4. Product-inhibition studies supported the postulated Hexa Uni Ping Pong (triple-transfer) reaction mechanism.
D002846:Chromatography, Affinity; D002847:Chromatography, Agarose; D007700:Kinetics; D008744:Methylamines; D010105:Oxygenases / Q000302:isolation & purification / Q000378:metabolism*; D011549:Pseudomonas / Q000201:enzymology*; D012685:Sepharose / Q000031:analogs & derivatives
D008744:Methylamines; D012685:Sepharose; D010105:Oxygenases