Carboxyl-terminal hydrophilic tail of a NhaP type Na +/H + antiporter from cyanobacteria is involved in the apparent affinity for Na + and pH sensitivity

Abstract

Little information is available on the C-terminal hydrophilic tails of prokaryotic Na +/H + antiporters. To address functional properties of the C-terminal tail, truncation mutants in this domain were constructed. Truncation of C-terminal amino acid residues of NhaP1 type antiporter from Synechocystis PCC6803 (SynNhaP1) did not change the V max values, but increased the K m values for Na + and Li + about 3 to 15-fold. Truncation of C-terminal tail of a halotolerant cyanobacterium Aphanothece halophytica (ApNhaP1) significantly decreased the V max although it did not alter the K m values for Na +. The C-terminal part of SynNhaP1 was expressed in E. coli and purified as a 16 kDa soluble protein. Addition of purified polypeptide to the membrane vesicles expressing the C-terminal truncated SynNhaP1 increased the exchange activities. Change of Glu519 and Glu521 to Lys in C-terminal tail altered the pH dependence of Na +/H + and Li +/H + exchange activities. These results indicate that the specific acidic amino acid residues at C-terminal domain play important roles for the K m and the pH dependence of the exchange activity.