Nitrate uptake by halotolerant cyanobacterium Aphanothece halophytica

Abstract

Nitrate uptake has been studied in the halotolerant cyanobacterium Aphanothece halophytica by monitoring nitrate remaining in the solution after contacting with the cells. Study of kinetics of nitrate uptake by cells grown under normal (0.5 M NaCl) and salt stress (2 M NaCl) conditions revealed Ks values of 416 and 450 micro-M respectively, the maximum velocities (Vmax) were 9.1 and 5.3 micro-mol/min/mgChl respectively. The ability of the cells to take up nitrate was effectively reduced by either ammonium addition or selective inhibition of CO2 fixation by DL-glyceraldehyde. Ammonium assimilation inhibitor, L-methionine sulfoximine was able to release nitrate uptake from ammonium inhibition. Overall results support the contention that nitrate utilization in cyanobacteria is regulated at the level of nitrate uptake through the concerted action of ammonium and CO2 fixation. Nitrite behaved as an effective competitive inhibitor of nitrate uptake, with a Ki (NO2-) of 84 micro-M. Nitrate and nitrite might be transported by the same carrier. Monensin depressed nitrate uptake in a concentration-dependent manner indicating that the nitrate uptake relies on the maintenance of a Na+ electrochemical gradient. Amiloride, a potent inhibitor of Na+/H+ antiport and Na+ channels, caused a decreased rate of nitrate uptake suggesting that nitrate uptake in A. halophytica was partly due to the Na+/H+ antiport.N,N-dicyclohexylcarbodiimide and carbonylcyanidetrifluoromethoxyphenyl-hydrazone are potent inhibitors of nitrate uptake in Aphanothece halophytica. These results might indicate that a pH gradient generated by H+/ATPase drives nitrate uptake in Aphanothece halophytica.