The ammonia transport, retention and futile cycling problem in cyanobacteria.

Ammonia is the preferred nitrogen source for many algae including the cyanobacterium Synechococcus elongatis (Synechococcus R-2; PCC 7942). Modelling ammonia uptake by cells is not straightforward because it exists in solution as NH(3) and NH (4) (+) . NH(3) is readily diffusible not only via the lipid bilayer but also through aquaporins and other more specific porins. On the other hand, NH (4) (+) requires cationic transporters to cross a membrane. Significant intracellular ammonia pools (≈1-10 mol m(-3)) are essential for the synthesis of amino acids from ammonia. The most common model envisaged for how cells take up ammonia and use it as a nitrogen source is the "pump-leak model" where uptake occurs through a simple diffusion of NH(3) or through an energy-driven NH (4) (+) pump balancing a leak of NH(3) out of the cell. The flaw in such models is that cells maintain intracellular pools of ammonia much higher than predicted by such models. With caution, [(14)C]-methylamine can be used as an analogue tracer for ammonia and has been used to test various models of ammonia transport and metabolism. In this study, simple "proton trapping" accumulation by the diffusion of uncharged CH(3)NH(2) has been compared to systems where CH(3)NH (3) (+) is taken up through channels, driven by the membrane potential (ΔU (i,o)) or the electrochemical potential for Na(+) (ΔμNa (i,o) (+) ). No model can be reconciled with experimental data unless the permeability of CH(3)NH(2) across the cell membrane is asymmetric: permeability into the cell is very high through gated porins, whereas permeability out of the cell is very low (≈40 nm s(-1)) and independent of the extracellular pH. The best model is a Na (in) (+) /CH(3)NH (3) (+) (in) co-porter driven by ΔμNa (i,o) (+) balancing synthesis of methylglutamine and a slow leak governed by Ficks law, and so there is significant futile cycling of methylamine across the cell membrane to maintain intracellular methylamine pools high enough for fixation by glutamine synthetase. The modified pump-leak model with asymmetric permeability of the uncharged form is a viable model for understanding ammonia uptake and retention in plants, free-living microbes and organisms in symbiotic relationships.