METHYLAMMONIUM TRANSPORT IN ANACYSTIS-NIDULANS R-2

Methylammonium was taken up rapidly by illuminated cells of A. nidulans R-2, leading to internal concentrations of 1.3 .+-. 0.1 mM within 1 min, and a gradient of up to 200 between the cells and medium. Accumulation of 14CH3NH3+ required at least 5 mM NaCl, but the uptake rate was independent of medium pH between 6.5 and 9. The kinetics of uptake could be resolved into an initial fast phase lasting < 1 min (approximate Km, 7.2 .mu.M; Vmax, 12.5 nmol min-1 mg of protein-1 at 15.degree. C). A 2nd, slower phase associated with product formation was eliminated by preincubation with methionine sulfoximine, a specific inhibitor of glutamine synthetase; the rapid phase was unaffected by this treatment. Ammonium ions competed with 14CH3NH3+ for entry, and addition of 5 .mu.M NH4+ or 100 .mu.M CH3NH3+ released 14CH3NH3+ accumulated during the rapid phase of entry. Small additions of NH4+ made at the same time as additions of 14CH3NH3+ delayed the start of radioactivity uptake by a time which corresponded accurately with the period needed for the complete removal of the added NH4+. The effects of inhibitors on accumulation and carbocyanine dye fluorescence suggest that ATP-dependent membrane potential was needed to drive 14CH3NH3+ transport. Spheroplasts were as active as whole cells in accumulating NH4+ and 14CH3NH3+, indicating that soluble periplasmic components are not involved in the translocation. Some significant differences between the translocation of 14CH3NH3+ and that of NH4+ were observed: growth with NH4+ in place of NO3- repressed 14CH3NH3+ accumulation ability without affecting the NH4+ uptake rate, Na+ was not required for NH4+ uptake and concentrations of KCl inhibitory with 14CH3NH3+ did not reduce NH4+ uptake.