Binding of monovalent cations to methylamine dehydrogenase in the semiquinone state and its effect on electron transfer.

The binding of monovalent cations to methylamine dehydrogenase in the semiquinone state (MADHsq) at a site close to the tryptophan tryptophylquinone (TTQ) active center is demonstrated in experiments which show that the radical EPR signal of MADHsq is considerably broadened in the presence of Cs+, NH4+, and, to a smaller extent, Na+. The cations also stabilize the semiquinone state, as is evident from the increase of the EPR intensity they induce. On the basis of the optical absorbance spectra, two slightly different forms of MADHsq can be discerned. One form, with the main band at 425 nm, is observed at low pH and in the presence of NH4+, whereas the other, with the main band at 429 nm, is observed at high pH and in the presence of Cs+ or Na+. Stopped-flow studies of the oxidation by amicyanin of MADHred via MADHsq to MADHox show a strong stimulation of the first step by monovalent cations. It is shown that it is primarily the actual electron transfer rate, rather than the affinity of MADHred for amicyanin, that is affected by cations. Values for the dissociation constants of the monovalent cations for MADHred, estimated from the kinetic experiments, are higher than those that were previously determined for MADHox, and can be deduced to be higher than those for MADHsq as well. The results are discussed within the context of the electron transfer theory.