Reactions of benzylamines with methylamine dehydrogenase. Evidence for a carbanionic reaction intermediate and reaction mechanism similar to eukaryotic quinoproteins
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 Ki 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 Ks 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 Ks and sigma p. Plots of log k3 and log Ks 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 Ks was primarily influenced by resonance effects. No correlation was observed between either k3 or Ks and the relative hydrophobicity of the para-substituted benzylamines or steric parameters.(ABSTRACT TRUNCATED AT 250 WORDS)