N-Methylglutamate Dehydrogenase, a Flavohaemoprotein Purified from a New Pink Trimethylamine-utilizing Bacterium

Abstract

A new strain of pink facultative methylotroph, bacterium AT2, capable of growth on methylamine, trimethylamine, methanol, formate and a range of non-C1 substrates has been isolated. On the basis of the enzyme activities in cell-free extracts, the organism appears to have the same pathway for trimethylamine oxidation as Pseudomonas aminovorans, i.e. via trimethylamine N-oxide and N-methylglutamate. N-Methylglutamate dehydrogenase in this organism was a ‘soluble’ enzyme (i.e. was not sedimented at 100000 g in 60 min) and reacted with the electron acceptors phenazine methosulphate, 2,6-dichlorophenolindophenol, Wurster’s blue and the radical cation of 2,2′-azinodi-[3-ethylbenzthiazoline 6-sulphonate]. It was not active with NAD, ferricyanide or cytochrome c. The enzyme was purified to apparent homogeneity (as assessed by polyacrylamide gel electrophoresis) and was shown to contain flavin and cytochrome c, both of which could be reduced by N-methylglutamate. The flavin prosthetic group could be liberated by boiling and was probably FAD. The true K m of the enzyme for N-methylglutamate was 0·33 mm and for 2,6-dichlorophenolindophenol, 0·20 mm.