The C-terminal cytoplasmic portion of the NhaP2 cation-proton antiporter from Vibrio cholerae affects its activity and substrate affinity

作者：Wiens Evan J; Winogrodzki Judith L; Resch Craig T; Orriss George L; Stetefeld Joerg; Dibrov Pavel^*

来源：Molecular and Cellular Biochemistry, 2014, 389(1-2): 51-58.

DOI：10.1007/s11010-013-1926-x

• Soluble domain
• Deletion analysis
• cation/proton antiporter
• Vibrio cholerae

In this work, we report the phenotypic and biochemical effects of deleting the C-terminal cytoplasmic portion of the NhaP2 cation/proton antiporter from Vibrio cholerae. While the deletion changed neither the expression nor targeting of the Vc-NhaP2 in an antiporter-less Escherichia coli strain, it resulted in a changed sensitivity of the host to sodium ions at neutral pH, indicating an altered Na+ transport through the truncated variant. When assayed in inside-out sub-bacterial vesicles, the truncation was found to result in greatly reduced K+/H+ and Na+/H+ antiport activity at all pH values tested and a greater than fivefold decrease in the affinity for K+ (measured as the apparent K (m)) at pH 7.5. Being expressed in trans in a strain of V. cholerae bearing a chromosomal nhaP2 deletion, the truncated nhaP2 gene was able to complement its inability to grow in potassium-rich medium at pH 6.0. Thus the residual K+/H+ antiport activity associated with the truncated Vc-NhaP2 was still sufficient to protect cells from an over-accumulation of K+ ions in the cytoplasm. The presented data suggest that while the cytoplasmic portion of Vc-NhaP2 is not involved in ion translocation directly, it is necessary for optimal activity and substrate binding of the Vc-NhaP2 antiporter.

• 出版日期2014-4