Cloning, functional expression in Escherichia coli and primary characterization of a new Na+/H+ antiporter, NhaD, of Vibrio cholerae.
Vibrio cholerae is the infectious agent of the deadly diarrheal disease, cholera. Na+ ion homeostasis is believed to play a key role in both physiology and pathogenicity of this bacterium. However, molecular mechanisms of sodium exchange in V. cholerae are still poorly understood. In the present work a gene encoding an unusual Na+/H+ antiporter, nhaD, was identified in the V. cholerae genome. nhaD was cloned from Vibrio cholerae and expressed in Escherichia coli. The antiporter functioned in an E. coli nhaAnhaB mutant strain to confer resistance to LiCl and NaCl. When assayed in inside-out subbacterial vesicles, V. cholerae NhaD demonstrated high affinity for Na+ ions (1.1 mM Na+ was required for the half-maximal response at the pH-optimum). The most striking feature of Vc-NhaD is a unique pH-profile of its activity with a sharp maximum at pH 8.0, different from that of any bacterial sodium-proton antiporter described so far. The difference is rationalized as being the result of a His to Arg substitution in a putative pH sensing residue.
Dzioba
Judith
J
Department of Microbiology, Faculty of Science, University of Manitoba, Winnipeg, Canada.
Ostroumov
Elena
E
Winogrodzki
Arthur
A
Dibrov
Pavel
P
D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D018021:Lithium Chloride / Q000378:metabolism; D012965:Sodium Chloride / Q000378:metabolism; D017923:Sodium-Hydrogen Exchangers / Q000235:genetics* / Q000378:metabolism; D014734:Vibrio cholerae / Q000235:genetics* / Q000378:metabolism
D001426:Bacterial Proteins; C111423:NhaD protein, Vibrio; D017923:Sodium-Hydrogen Exchangers; D012965:Sodium Chloride; D018021:Lithium Chloride