| Abstract |
Ohyama et al. previously isolated Escherichia coli mutant RS1, which had a negligible activity for sodium ion extrusion at alkaline pH (T. Ohyama, R. Imaizumi, K. Igarashi, and H. Kobayashi, J. Bacteriol. 174:7743-7749, 1992). Our present study showed that the mutation of RS1 was compensated for by a cloned chaA gene. It has been proposed that sodium ion extrusion by ChaA is prevented under physiological conditions (D. M. Ivey, A. A. Guffanti, J. Zemsky, E. Pinner, R. Karpel, E. Padan, S. Schuldiner, and T. A. Krulwich, J. Biol. Chem. 268:11296-11303, 1993). In order to clarify the physiological role of chaA in sodium ion circulation at alkaline pH, we constructed a delta chaA mutant. The resultant mutant, TO112, deficient in both nhaA and chaA, was unable to grow at pH 8.5 in medium containing 0.1 M sodium chloride and had negligible sodium ion extrusion activity. However, TO112 grew at pH 7.0 in medium containing 0.4 M sodium chloride. Sodium ions were extruded from TO112 cells at neutral pH. The extrusion activity at pH 7.5 was greatly reduced by the deletion of nhaB. These data demonstrate that the activity of nhaB is low at high pH and that ChaA extrudes sodium ions at alkaline pH. The uptake of calcium ions by everted membrane vesicles prepared from the delta chaA mutant TO110 was 60% of the activity observed in the vesicles of the wild-type strain at pH 8.5, but the activity at neutral pH was not reduced by the deletion of chaA. Therefore, it was also suggested that ChaA plays a role in calcium ion circulation at alkaline pH.
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