The Vibrio cholerae Mrp System: Cation/Proton Antiport Properties and Enhancement of Bile Salt Resistance in a Heterologous Host

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Abstract
The mrp operon from Vibrio cholerae encoding a putative multisubunit Na+/H+ antiporter was cloned and functionally expressed in the antiporter-deficient strain of Escherichia coli EP432. Cells of EP432 expressing Vc-Mrp exhibited resistance to Na+ and Li+ as well as to natural bile salts such as sodium cholate and taurocholate. When assayed in everted membrane vesicles of the E. coli EP432 host, Vc-Mrp had sufficiently high antiport activity to facilitate the first extensive analysis of Mrp system from a Gram-negative bacterium encoded by a group 2 mrp operon. Vc-Mrp was found to exchange protons for Li+, Na+, and K+ ions in pH-dependent manner with maximal activity at pH 9.0–9.5. Exchange was electrogenic (more than one H+ translocated per cation moved in opposite direction). The apparent Km at pH 9.0 was 1.08, 1.30, and 68.5 mM for Li+, Na+, and K+, respectively. Kinetic analyses suggested that Vc-Mrp operates in a binding exchange mode with all cations and protons competing for binding to the antiporter. The robust ion antiport activity of Vc-Mrp in sub-bacterial vesicles and its effect on bile resistance of the heterologous host make Vc-Mrp an attractive experimental model for the further studies of biochemistry and physiology of Mrp systems.