[5] N-Methyl-L-glutamate synthase

N-Methyl-L-glutamate synthase is an inducible enzyme found in several species of Pseudomonas grown on methylated amines. The enzyme functions in a cycle in which free methylamine first reacts with L-glutamate to yield N-methyl-L-glutamate and free ammonia. The N-methyl-L-glutamate is then oxidized to release formaldehyde and regenerate L-glutamate. This cycle provides both ammonia and an activated C-1 compound for growth of the microorganism. The reaction between L-glutamate and methylamine is measured by following the incorporation of [¹⁴C]methylamine into N-methyl-L-glutamate. Free methylamine is separated from N-methyl-L-glutamate by chromatography on Dowex 50-H⁺. The reaction is initiated by the addition of an appropriate amount of enzyme and incubated at 37^° for 15–30 min. Termination of the reaction is accomplished by the addition of 50 μl of 20% trichloroacetic acid, and the denatured protein is removed by centrifugation.