Functional expression and initial biochemical characterization of Yp-NhaP, cation-proton antiporter from Yersinia pestis
Date
2011-09-06
Authors
Abboud, Talal
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Abstract
The major objectives of this work were cloning, functional expression and primary biochemical characterization of Yp-NhaP, putative sodium-proton antiporter from the dangerous human pathogen Yersinia pestis.
We expressed Yp-NhaP in its functional form in the antiporter-deficient strain of E. coli, TO114. When assayed in inside-out sub-bacterial membrane vesicles, Yp-NhaP acted as an electroneutral cation/proton antiporter, exchanging Ca2+, K+, Na+ and Li+ ions for H+.
Competition experiments suggested that in vivo Yp-NhaP operates as Ca2+/H+ and, possibly, Ca2+/Na+ antiporter rather than K+/H+ or Na+/H+ antiporter. Ca2+/H+ and Li+/H+ antiport catalyzed by Yp-NhaP peaked at pH close to 8.0, while K+/H+ and Na+/H+ antiport were smoothly increasing from pH 6.5 to pH 9.0. We also observed inhibition by the excess of substrate in the case of Ca2+/H+ and Li+/H+ antiport mediated by Yp-NhaP.
As expected, chromosomal deletion of Yp-nhaP gene did not affect resistance of Y. pestis cells to alkali cations.
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Keywords
Y.pestis, Antiporter