1887

Microbiology Society logo

Volume 152, Issue 11

The Phn system of : a second high-affinity ABC-transporter for phosphate Free

Abstract

Uptake of inorganic phosphate, an essential but often limiting nutrient, in bacteria is usually accomplished by the high-affinity ABC-transport system Pst. Pathogenic species of mycobacteria contain several copies of the genes encoding the Pst system (), and two of the encoded proteins, PstS1 and PstS2, have been shown to be virulence factors in . The fast-growing contains only a single copy of the operon. This study reports the biochemical and molecular characterization of a second high-affinity phosphate transport system, designated Phn. The Phn system is encoded by a three-gene operon that constitutes the components of a putative ABC-type phosphonate/phosphate transport system. Expression studies using and transcriptional fusions showed that both operons were induced when the culture entered phosphate limitation, indicating a role for both systems in phosphate uptake at low extracellular concentrations. Deletion mutants in either or failed to grow in minimal medium with a 10 mM phosphate concentration, while the isogenic wild-type strain mc155 grew at micromolar phosphate concentrations. Analysis of the kinetics of phosphate transport in the wild-type and mutant strains led to the proposal that the Phn and Pst systems are both high-affinity phosphate transporters with similar affinities for phosphate (i.e. apparent values between 40 and 90 μM P). The Phn system of appears to be unique in that, unlike previously identified Phn systems, it does not recognize phosphonates or phosphite as substrates.

  • Received:
  • Accepted:
  • Revised:
  • Published Online:
Keyword(s): ABC, ATP-binding cassette , MU, Miller units , Pit, phosphate inorganic transport and Pst, phosphate-specific transport
SGM
Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.29201-0
2006-11-01
2024-04-25
Loading full text...

Full text loading...

/deliver/fulltext/micro/152/11/3453.html?itemId=/content/journal/micro/10.1099/mic.0.29201-0&mimeType=html&fmt=ahah

References

  1. Alexeyev M. F, Shokolenko I. N, Croughan T. P. 1995; Improved antibiotic-resistance gene cassettes and omega elements for Escherichia coli vector construction and in vitro deletion/insertion mutagenesis. Gene 160:63–67 [CrossRef]
     [Google Scholar]
  2. Allenby N. E, O'Connor N, Pragai Z, Carter N. M, Miethke M, Engelmann S, Wipat A, Ward A. C, Harwood C. R. 2004; Post-transcriptional regulation of the Bacillus subtilis pst operon encoding a phosphate-specific ABC transporter. Microbiology 150:2619–2628 [CrossRef]
     [Google Scholar]
  3. Allenby N. E, O'Connor N, Pragai Z, Ward A. C, Wipat A, Harwood C. R. 2005; Genome-wide transcriptional analysis of the phosphate starvation stimulon of Bacillus subtilis . J Bacteriol 187:8063–8080 [CrossRef]
     [Google Scholar]
  4. Bardin S. D, Finan T. M. 1998; Regulation of phosphate assimilation in Rhizobium (Sinorhizobium) meliloti . Genetics 148:1689–1700
     [Google Scholar]
  5. Bardin S, Dan S, Osteras M, Finan T. M. 1996; A phosphate transport system is required for symbiotic nitrogen fixation by Rhizobium meliloti . J Bacteriol 178:4540–4547
     [Google Scholar]
  6. Bardin S. D, Voegele R. T, Finan T. M. 1998; Phosphate assimilation in Rhizobium (Sinorhizobium) meliloti : identification of a pit -like gene. J Bacteriol 180:4219–4226
     [Google Scholar]
  7. Bhatt K, Banerjee S. K, Chakraborti P. K. 2000; Evidence that phosphate specific transporter is amplified in a fluoroquinolone resistant Mycobacterium smegmatis . Eur J Biochem 267:4028–4032 [CrossRef]
     [Google Scholar]
  8. Braibant M, Ooms J, Peirs P, Huygen K, Wattiez R, Content J, Lefèvre P, de Wit L. 1996a; Identification of a second Mycobacterium tuberculosis gene cluster encoding proteins of an ABC phosphate transporter. FEBS Lett 394:206–212 [CrossRef]
     [Google Scholar]
  9. Braibant M, Peirs P, Ooms J, Huygen K, Andersen A. B, Content J, Lefèvre P, de Wit L. 1996b; A Mycobacterium tuberculosis gene cluster encoding proteins of a phosphate transporter homologous to the Escherichia coli Pst system. Gene 176:171–176 [CrossRef]
     [Google Scholar]
  10. Chen C. M, Ye Q. Z, Zhu Z. M, Wanner B. L, Walsh C. T. 1990; Molecular biology of carbon-phosphorus bond cleavage. Cloning and sequencing of the phn (psiD) genes involved in alkylphosphonate uptake and C–P lyase activity in Escherichia coli B. J Biol Chem 265:4461–4471
     [Google Scholar]
  11. Collins D. M, Kawakami R. P, Buddle B. M, Wards B. J, de Lisle G. W. 2003; Different susceptibility of two animal species infected with isogenic mutants of Mycobacterium bovis identifies phoT as having roles in tuberculosis virulence and phosphate transport. Microbiology 149:3203–3212 [CrossRef]
     [Google Scholar]
  12. Gonzalez-y-Merchand J. A, Estrada-Garcia I, Colston M. J, Cox R. A. 1996; A novel method for the isolation of mycobacterial DNA. FEMS Microbiol Lett 135:71–77 [CrossRef]
     [Google Scholar]
  13. Hanahan D, Jessee J, Bloom F. R. 1991; Plasmid transformation of Escherichia coli and other bacteria. Methods Enzymol 204:63–113
     [Google Scholar]
  14. Hoffer S. M, Schoondermark P, van Veen H. W, Tommassen J. 2001; Activation by gene amplification of pitB , encoding a third phosphate transporter of Escherichia coli K-12. J Bacteriol 183:4659–4663 [CrossRef]
     [Google Scholar]
  15. Hulett F. M. 1995; Complex phosphate regulation by sequential switches in Bacillus subtilis. In Two-Component Signal Transduction pp  298–302 Edited by Hoch J. A., Silhavy T. J. Washington, DC: American Society for Microbiology;
     [Google Scholar]
  16. Hulett F. M, Jensen K. 1988; Critical roles of spo0A and spo0H in vegetative alkaline phosphatase production in Bacillus subtilis . J Bacteriol 170:3765–3768
     [Google Scholar]
  17. Hulett F. M, Lee J, Shi L, Sun G, Chesnut R, Sharkova E, Duggan M. F, Kapp N. 1994; Sequential action of two-component genetic switches regulates the PHO regulon in Bacillus subtilis . J Bacteriol 176:1348–1358
     [Google Scholar]
  18. Imazu K, Tanaka S, Kuroda A, Anbe Y, Kato J, Ohtake H. 1998; Enhanced utilization of phosphonate and phosphite by Klebsiella aerogenes . Appl Environ Microbiol 64:3754–3758
     [Google Scholar]
  19. Kononova S. V, Nesmeyanova M. A. 2002; Phosphonates and their degradation by microorganisms. Biochemistry 67:184–195
     [Google Scholar]
  20. Kriakov J, Lee S, Jacobs W. R. Jr 2003; Identification of a regulated alkaline phosphatase, a cell surface-associated lipoprotein, in Mycobacterium smegmatis . J Bacteriol 185:4983–4991 [CrossRef]
     [Google Scholar]
  21. Lee M. H, Pascopella L, Hatfull G. F, Jacobs W. R. Jr 1991; Site-specific integration of mycobacteriophage L5: integration-proficient vectors for Mycobacterium smegmatis , Mycobacterium tuberculosis , and bacille Calmette-Guerin. Proc Natl Acad Sci U S A 88:3111–3115 [CrossRef]
     [Google Scholar]
  22. Lefèvre P, Braibant M, de Wit L, Kalai M, Roeper D, Grotzinger J, Delville J. P, Peirs P, Ooms J, Huygen K, Content J. 1997; Three different putative phosphate transport receptors are encoded by the Mycobacterium tuberculosis genome and are present at the surface of Mycobacterium bovis BCG. J Bacteriol 179:2900–2906
     [Google Scholar]
  23. Licha I, Benes I, Janda S, Host'alek Z, Janacek K. 1997; Characterization of phosphate transport in Streptomyces granaticolor . Biochem Mol Biol Int 41:431–437
     [Google Scholar]
  24. McMullan G, Quinn J. P. 1994; In vitro characterization of a phosphate starvation-independent carbon-phosphorus bond cleavage activity in Pseudomonas fluorescens 23F. J Bacteriol 176:320–324
     [Google Scholar]
  25. Menard R, Sansonetti P. J, Parsot C. 1993; Nonpolar mutagenesis of the ipa genes defines IpaB, IpaC, and IpaD as effectors of Shigella flexneri entry into epithelial cells. J Bacteriol 175:5899–5906
     [Google Scholar]
  26. Metcalf W. W, Wanner B. L. 1991; Involvement of the Escherichia coli phn (psiD) gene cluster in assimilation of phosphorus in the form of phosphonates, phosphite, P[sub]i[/sub] esters, and P[sub]i[/sub]. J Bacteriol 173:587–600
     [Google Scholar]
  27. Metcalf W. W, Wolfe R. S. 1998; Molecular genetic analysis of phosphite and hypophosphite oxidation by Pseudomonas stutzeri WM88. J Bacteriol 180:5547–5558
     [Google Scholar]
  28. Meyers P. R, Bourn W. R, Steyn L. M, Beyers A. D, Brown G. D, van Helden P. D. 1998; Novel method for rapid measurement of growth of mycobacteria in detergent-free media. J Clin Microbiol 36:2752–2754
     [Google Scholar]
  29. Monk B. C, Kurtz M. B, Marrinan J. A, Perlin D. S. 1991; Cloning and characterization of the plasma membrane H[sup]+[/sup]-ATPase from Candida albicans . J Bacteriol 173:6826–6836
     [Google Scholar]
  30. Obojska A, Lejczak B, Kubrak M. 1999; Degradation of phosphonates by streptomycete isolates. Appl Microbiol Biotechnol 51:872–876 [CrossRef]
     [Google Scholar]
  31. Peirs P, Boarbi S, Wang X. M, Denis O, Braibant M, Pethe K, Locht C, Huygen K, Content J, Lefèvre P. 2005; Mycobacterium tuberculosis with disruption in genes encoding the phosphate binding proteins PstS1 and PstS2 is deficient in phosphate uptake and demonstrates reduced in vivo virulence. Infect Immun 73:1898–1902 [CrossRef]
     [Google Scholar]
  32. Pelicic V, Jackson M, Reyrat J. M, Gicquel B, Guilhot C, Jacobs W. R. Jr 1997; Efficient allelic exchange and transposon mutagenesis in Mycobacterium tuberculosis . Proc Natl Acad Sci U S A 94:10955–10960 [CrossRef]
     [Google Scholar]
  33. Perez E, Samper S, Bordas Y, Guilhot C, Gicquel B, Martin C. 2001; An essential role for phoP in Mycobacterium tuberculosis virulence. Mol Microbiol 41:179–187 [CrossRef]
     [Google Scholar]
  34. Pirt S. J. 1975 Principles of Microbe and Cell Cultivation Oxford: Blackwell Scientific Publications;
     [Google Scholar]
  35. Qi Y, Kobayashi Y, Hulett F. M. 1997; The pst operon of Bacillus subtilis has a phosphate-regulated promoter and is involved in phosphate transport but not in regulation of the Pho regulon. J Bacteriol 179:2534–2539
     [Google Scholar]
  36. Rao M, Streur T. L, Aldwell F. E, Cook G. M. 2001; Intracellular pH regulation by Mycobacterium smegmatis and Mycobacterium bovis BCG. Microbiology 147:1017–1024
     [Google Scholar]
  37. Ruiz N, Silhavy T. J. 2003; Constitutive activation of the Escherichia coli Pho regulon upregulates rpoS translation in an Hfq-dependent fashion. J Bacteriol 185:5984–5992 [CrossRef]
     [Google Scholar]
  38. Sambrook J, Fritsch E. F, Maniatis T. 1989 Molecular Cloning: a Laboratory Manual, 2nd edn.. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
     [Google Scholar]
  39. Snapper S. B, Melton R. E, Mustafa S, Kieser T, Jacobs W. R. Jr 1990; Isolation and characterization of efficient plasmid transformation mutants of Mycobacterium smegmatis . Mol Microbiol 4:1911–1919 [CrossRef]
     [Google Scholar]
  40. Sola-Landa A, Moura R. S, Martin J. F. 2003; The two-component PhoR-PhoP system controls both primary metabolism and secondary metabolite biosynthesis in Streptomyces lividans . Proc Natl Acad Sci U S A 100:6133–6138 [CrossRef]
     [Google Scholar]
  41. Sola-Landa A, Rodriguez-Garcia A, Franco-Dominguez E, Martin J. F. 2005; Binding of PhoP to promoters of phosphate-regulated genes in Streptomyces coelicolor : identification of PHO boxes. Mol Microbiol 56:1373–1385 [CrossRef]
     [Google Scholar]
  42. Timm J, Lim E. M, Gicquel B. 1994; Escherichia coli -mycobacteria shuttle vectors for operon and gene fusions to lacZ : the pJEM series. J Bacteriol 176:6749–6753
     [Google Scholar]
  43. Tran S. L, Rao M, Simmers C, Gebhard S, Olsson K, Cook G. M. 2005; Mutants of Mycobacterium smegmatis unable to grow at acidic pH in the presence of the protonophore carbonyl cyanide m-chlorophenylhydrazone. Microbiology 151:665–672 [CrossRef]
     [Google Scholar]
  44. van Veen H. W. 1997; Phosphate transport in prokaryotes: molecules, mediators and mechanisms. Antonie Van Leeuwenhoek 72:299–315 [CrossRef]
     [Google Scholar]
  45. van Veen H. W, Abee T, Kortstee G. J, Konings W. N, Zehnder A. J. 1993; Mechanism and energetics of the secondary phosphate transport system of Acinetobacter johnsonii 210A. J Biol Chem 268:19377–19383
     [Google Scholar]
  46. van Veen H. W, Abee T, Kortstee G. J, Konings W. N, Zehnder A. J. 1994a; Substrate specificity of the two phosphate transport systems of Acinetobacter johnsonii 210A in relation to phosphate speciation in its aquatic environment. J Biol Chem 269:16212–16216
     [Google Scholar]
  47. van Veen H. W, Abee T, Kortstee G. J, Konings W. N, Zehnder A. J. 1994b; Translocation of metal phosphate via the phosphate inorganic transport system of Escherichia coli . Biochemistry 33:1766–1770 [CrossRef]
     [Google Scholar]
  48. Voegele R. T, Bardin S, Finan T. M. 1997; Characterization of the Rhizobium (Sinorhizobium) meliloti high- and low-affinity phosphate uptake systems. J Bacteriol 179:7226–7232
     [Google Scholar]
  49. von Kruger W. M, Humphreys S, Ketley J. M. 1999; A role for the PhoBR regulatory system homologue in the Vibrio cholerae phosphate-limitation response and intestinal colonization. Microbiology 145:2463–2475
     [Google Scholar]
  50. Wanner B. L. others 1996; Phosphorus assimilation and control of the phosphate regulon. In Escherichia coli and Salmonella: Cellular and Molecular Biology pp.  1357–1381 Edited by Neidhardt F. C. Washington, DC: American Society for Microbiology;
     [Google Scholar]
  51. Wanner B. L, Chang B. D. 1987; The phoBR operon in Escherichia coli K-12. J Bacteriol 169:5569–5574
     [Google Scholar]
  52. White A. K, Metcalf W. W. 2004a; The htx and ptx operons of Pseudomonas stutzeri WM88 are new members of the Pho regulon. J Bacteriol 186:5876–5882 [CrossRef]
     [Google Scholar]
  53. White A. K, Metcalf W. W. 2004b; Two C-P lyase operons in Pseudomonas stutzeri and their roles in the oxidation of phosphonates, phosphite, and hypophosphite. J Bacteriol 186:4730–4739 [CrossRef]
     [Google Scholar]
  54. Willsky G. R, Malamy M. H. 1980; Characterization of two genetically separable inorganic phosphate transport systems in Escherichia coli . J Bacteriol 144:356–365
     [Google Scholar]
  55. Yuan Z. C, Zaheer R, Finan T. M. 2006; Regulation and properties of PstSCAB, a high-affinity, high-velocity phosphate transport system of Sinorhizobium meliloti . J Bacteriol 188:1089–1102 [CrossRef]
     [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.29201-0
Loading
The Phn system of Mycobacterium smegmatis: a second high-affinity ABC-transporter for phosphate
Microbiology 152, 3453 (2006); https://doi.org/10.1099/mic.0.29201-0
/content/journal/micro/10.1099/mic.0.29201-0
/content/journal/micro/10.1099/mic.0.29201-0
Loading

Data & Media loading...

Most cited Most Cited RSS feed

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