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Title: Functional expression and initial biochemical characterization of Yp-NhaP, cation-proton antiporter from Yersinia pestis
Authors: Abboud, Talal
Supervisor: Dibrov, Pavel (Microbiology)
Examining Committee: Court, Deborah (Microbiology) Khajehpour, Mazdak (Chemistry)
Graduation Date: October 2011
Keywords: Y.pestis
Issue Date: 6-Sep-2011
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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