1887

Abstract

has adapted to a wide range of salinity, pH and osmotic conditions, enabling it to survive passage through the host and persist in the environment. Among the many proteins responsible for bacterial survival under these diverse conditions, we have identified Vc-NhaP1 as a K(Na)/H antiporter essential for growth at low environmental pH. Deletion of the gene caused growth inhibition when external potassium was either limited (100 mM and below) or in excess (400 mM and above). This growth defect was most apparent at mid-exponential phase, after 4–6 h of culture. Using a pH-sensitive GFP, cytosolic pH was shown to be dependent on K in acidic external conditions in a Vc-NhaP1-dependent manner. When functionally expressed in an antiporterless strain and assayed in everted membrane vesicles, Vc-NhaP1 operated as an electroneutral alkali cation/proton antiporter, exchanging K or Na ions for H within a broad pH range (7.25–9.0). These data establish the putative NhaP1 protein as a functional K(Na)/H antiporter of the CPA1 family that is required for bacterial pH homeostasis and growth in an acidic environment.

Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.056119-0
2012-04-01
2024-04-26
Loading full text...

Full text loading...

/deliver/fulltext/micro/158/4/1094.html?itemId=/content/journal/micro/10.1099/mic.0.056119-0&mimeType=html&fmt=ahah

References

  1. Bakker E. P., Mangerich W. E. ( 1981). Interconversion of components of the bacterial proton motive force by electrogenic potassium transport. J Bacteriol 147:820–826[PubMed]
    [Google Scholar]
  2. Corratgé-Faillie C., Jabnoune M., Zimmermann S., Véry A. A., Fizames C., Sentenac H. ( 2010). Potassium and sodium transport in non-animal cells: the Trk/Ktr/HKT transporter family. Cell Mol Life Sci 67:2511–2532 [View Article][PubMed]
    [Google Scholar]
  3. Csonka L. N. ( 1989). Physiological and genetic responses of bacteria to osmotic stress. Microbiol Rev 53:121–147[PubMed]
    [Google Scholar]
  4. Dzioba J., Ostroumov E., Winogrodzki A., Dibrov P. ( 2002). Cloning, functional expression in Escherichia coli and primary characterization of a new Na+/H+ antiporter, NhaD, of Vibrio cholerae . Mol Cell Biochem 229:119–124 [View Article][PubMed]
    [Google Scholar]
  5. Dzioba-Winogrodzki J., Winogrodzki O., Krulwich T. A., Boin M. A., Häse C. C., Dibrov P. ( 2009). The Vibrio cholerae Mrp system: cation/proton antiport properties and enhancement of bile salt resistance in a heterologous host. J Mol Microbiol Biotechnol 16:176–186 [View Article][PubMed]
    [Google Scholar]
  6. Enserink M. ( 2010). Public health. No vaccines in the time of cholera. Science 329:1462–1463 [View Article][PubMed]
    [Google Scholar]
  7. Epstein W., Kim B. S. ( 1971). Potassium transport loci in Escherichia coli K-12. J Bacteriol 108:639–644[PubMed]
    [Google Scholar]
  8. Gómez-Puyou A., Gómez-Lojero C. ( 1977). The use of ionophores and channel formers in the study of the function of biological membranes. Current Topics in Bioenergetics vol. 6221–257 Rao Sanadi D. New York: Academic Press;
    [Google Scholar]
  9. Guffanti A. A., Cheng J., Krulwich T. A. ( 1998). Electrogenic antiport activities of the Gram-positive Tet proteins include a Na+(K+)/K+ mode that mediates net K+ uptake. J Biol Chem 273:26447–26454 [View Article][PubMed]
    [Google Scholar]
  10. Hamaide F., Kushner D. J., Sprott G. D. ( 1983). Proton motive force and Na+/H+ antiport in a moderate halophile. J Bacteriol 156:537–544[PubMed]
    [Google Scholar]
  11. Hamashima H., Iwasaki M., Arai T. ( 1995). A simple and rapid method for transformation of Vibrio species by electroporation. Methods Mol Biol 47:155–160[PubMed]
    [Google Scholar]
  12. Häse C. C., Fedorova N. D., Galperin M. Y., Dibrov P. A. ( 2001). Sodium ion cycle in bacterial pathogens: evidence from cross-genome comparisons. Microbiol Mol Biol Rev 65:353–370 [View Article][PubMed]
    [Google Scholar]
  13. Ho S. N., Hunt H. D., Horton R. M., Pullen J. K., Pease L. R. ( 1989). Site-directed mutagenesis by overlap extension using the polymerase chain reaction. Gene 77:51–59 [View Article][PubMed]
    [Google Scholar]
  14. Kitko R. D., Cleeton R. L., Armentrout E. I., Lee G. E., Noguchi K., Berkmen M. B., Jones B. D., Slonczewski J. L. ( 2009). Cytoplasmic acidification and the benzoate transcriptome in Bacillus subtilis . PLoS ONE 4:e8255 [View Article][PubMed]
    [Google Scholar]
  15. Kitko R. D., Wilks J. C., Garduque G. M., Slonczewski J. L. ( 2010). Osmolytes contribute to pH homeostasis of Escherichia coli . PLoS ONE 5:e10078 [View Article][PubMed]
    [Google Scholar]
  16. Kroll R. G., Booth I. R. ( 1981). The role of potassium transport in the generation of a pH gradient in Escherichia coli . Biochem J 198:691–698[PubMed]
    [Google Scholar]
  17. Kroll R. G., Booth I. R. ( 1983). The relationship between intracellular pH, the pH gradient and potassium transport in Escherichia coli . Biochem J 216:709–716[PubMed]
    [Google Scholar]
  18. Mekalanos J. J., Swartz D. J., Pearson G. D., Harford N., Groyne F., de Wilde M. ( 1983). Cholera toxin genes: nucleotide sequence, deletion analysis and vaccine development. Nature 306:551–557 [View Article][PubMed]
    [Google Scholar]
  19. Metcalf W. W., Jiang W., Daniels L. L., Kim S. K., Haldimann A., Wanner B. L. ( 1996). Conditionally replicative and conjugative plasmids carrying lacZα for cloning, mutagenesis, and allele replacement in bacteria. Plasmid 35:1–13 [View Article][PubMed]
    [Google Scholar]
  20. Miller C. J., Drasar B. S., Feachem R. G. ( 1984). Response of toxigenic Vibrio cholerae O1 to physico-chemical stresses in aquatic environments. J Hyg (Lond) 93:475–495 [View Article][PubMed]
    [Google Scholar]
  21. Nakamura T., Tokuda H., Unemoto T. ( 1982). Effects of pH and monovalent cations on the potassium ion exit from the marine bacterium, Vibrio alginolyticus, and the manipulation of cellular cation contents. Biomembranes 692:389–396 [View Article]
    [Google Scholar]
  22. Nakamura T., Tokuda H., Unemoto T. ( 1984). K+/H+ antiporter functions as a regulator of cytoplasmic pH in a marine bacterium, Vibrio alginolyticus . Biochim Biophys Acta 776:330–336 [View Article][PubMed]
    [Google Scholar]
  23. Nakamura T., Kawasaki S., Unemoto T. ( 1992). Roles of K+ and Na+ in pH homeostasis and growth of the marine bacterium Vibrio alginolyticus . J Gen Microbiol 138:1271–1276[PubMed] [CrossRef]
    [Google Scholar]
  24. Padan E., Zilberstein D., Schuldiner S. ( 1981). pH homeostasis in bacteria. Biochim Biophys Acta 650:151–166[PubMed] [CrossRef]
    [Google Scholar]
  25. Padan E., Maisler N., Taglicht D., Karpel R., Schuldiner S. ( 1989). Deletion of ant in Escherichia coli reveals its function in adaptation to high salinity and an alternative Na+/H+ antiporter system(s). J Biol Chem 264:20297–20302[PubMed]
    [Google Scholar]
  26. Radchenko M. V., Waditee R., Oshimi S., Fukuhara M., Takabe T., Nakamura T. ( 2006). Cloning, functional expression and primary characterization of Vibrio parahaemolyticus K+/H+ antiporter genes in Escherichia coli . Mol Microbiol 59:651–663 [View Article][PubMed]
    [Google Scholar]
  27. Resch C. T., Winogrodzki J. L., Patterson C. T., Lind E. J., Quinn M. J., Dibrov P., Häse C. C. ( 2010). The putative Na+/H+ antiporter of Vibrio cholerae, Vc-NhaP2, mediates the specific K+/H+ exchange in vivo. Biochemistry 49:2520–2528 [View Article][PubMed]
    [Google Scholar]
  28. Resch C. T., Winogrodzki J. L., Häse C. C., Dibrov P. ( 2011). Insights into the biochemistry of the ubiquitous NhaP family of cation/H+ antiporters. Biochem Cell Biol 89:130–137 [View Article][PubMed]
    [Google Scholar]
  29. Singleton F. L., Attwell R., Jangi S., Colwell R. R. ( 1982a). Effects of temperature and salinity on Vibrio cholerae growth. Appl Environ Microbiol 44:1047–1058[PubMed]
    [Google Scholar]
  30. Singleton F. L., Attwell R. W., Jangi M. S., Colwell R. R. ( 1982b). Influence of salinity and organic nutrient concentration on survival and growth of Vibrio cholerae in aquatic microcosms. Appl Environ Microbiol 43:1080–1085[PubMed]
    [Google Scholar]
  31. Slonczewski J. L., Rosen B. P., Alger J. R., Macnab R. M. ( 1981). pH homeostasis in Escherichia coli: measurement by 31P nuclear magnetic resonance of methylphosphonate and phosphate. Proc Natl Acad Sci U S A 78:6271–6275 [View Article][PubMed]
    [Google Scholar]
  32. Tokuda H., Unemoto T. ( 1981). A respiration-dependent primary sodium extrusion system functioning at alkaline pH in the marine bacterium Vibrio alginolyticus . Biochem Biophys Res Commun 102:265–271 [View Article][PubMed]
    [Google Scholar]
  33. Tokuda H., Unemoto T. ( 1982). Characterization of the respiration-dependent Na+ pump in the marine bacterium Vibrio alginolyticus . J Biol Chem 257:10007–10014[PubMed]
    [Google Scholar]
  34. Tokuda H., Nakamura T., Unemoto T. ( 1981). Potassium ion is required for the generation of pH-dependent membrane potential and delta pH by the marine bacterium Vibrio alginolyticus . Biochemistry 20:4198–4203 [View Article][PubMed]
    [Google Scholar]
  35. Vimont S., Berche P. ( 2000). NhaA, an Na+/H+ antiporter involved in environmental survival of Vibrio cholerae . J Bacteriol 182:2937–2944 [View Article][PubMed]
    [Google Scholar]
  36. Watten R. H., Morgan F. M., Songkhla Y. N., Vanikiati B., Phillips R. A. ( 1959). Water and electrolyte studies in cholera. J Clin Invest 38:1879–1889 [View Article][PubMed]
    [Google Scholar]
  37. Webb D. ( 1939). The sodium and potassium content of seawater. J Exp Biol 16:178–183
    [Google Scholar]
  38. Zhou W., Bertsova Y. V., Feng B., Tsatsos P., Verkhovskaya M. L., Gennis R. B., Bogachev A. V., Barquera B. ( 1999). Sequencing and preliminary characterization of the Na+-translocating NADH : ubiquinone oxidoreductase from Vibrio harveyi . Biochemistry 38:16246–16252 [View Article][PubMed]
    [Google Scholar]
  39. Zilberstein D., Agmon V., Schuldiner S., Padan E. ( 1982). The sodium/proton antiporter is part of the pH homeostasis mechanism in Escherichia coli . J Biol Chem 257:3687–3691[PubMed]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.056119-0
Loading
/content/journal/micro/10.1099/mic.0.056119-0
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error