It had been previously reported that aromatic amines were not substrates for the bacterial quinoprotein methylamine dehydrogenase. In this study, benzylamine-dependent activity was also not observed in the steady-state assay of this enzyme with the artificial electron acceptor phenazine ethosulfate (PES). Benzylamines did, however, stoichiometrically reduce the protein-bound tryptophan tryptophylquinone (TTQ) prosthetic group and acted as reversible competitive inhibitors of methylamine oxidation when the enzyme was assayed with PES. When methylamine dehydrogenase activity was monitored using a steady-state assay which employed its physiological electron acceptor amicyanin instead of PES, very low but detectable benzylamine-dependent activity was observed. The reactions of a series of para-substituted benzylamines with methylamine dehydrogenase were examined. A Hammett plot of the log of K(i) values for the competitive inhibition by these amines against sigma(p) exhibited a negative slope. Rapid kinetic measurements allowed the determination of values of k3 and K(s) for the reduction of TTQ by each of these amines. A Hammett plot of log k3 versus sigma(p) exhibited a positive slope, which suggests that the oxidation of these amines by methylamine dehydrogenase proceeds through a carbanionic reaction intermediate. A negative slope was observed for the correlation between log K(s) and sigma(p). Plots of log k3 and log K(s) against substituent constants which reflected either resonance or field/inductive parameters for each para substituent indicated that the magnitude of k3 was primarily influenced by field/inductive effects while K(s) was primarily influenced by resonance effects. No correlation was observed between either k3 Or K(s) and the relative hydrophobicity of the para-substituted benzylamines or steric parameters. The K(i) values which were obtained from steady-state kinetic experiments correlated strongly with the K(s) values which were obtained from rapid kinetic experiments. On the basis of these results, a mechanism is proposed for the reactions of benzylamines with this enzyme. These data are also discussed in light of results of similar studies of the reactions of para-substituted benzylamines with two eukaryotic quinoproteins, lysyl oxidase and plasma amine oxidase.