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. 1996 Aug;178(15):4628–4634. doi: 10.1128/jb.178.15.4628-4634.1996

Organization, transcription, and expression of the 5' region of the fla operon of Treponema phagedenis and Treponema pallidum.

R J Limberger 1, L L Slivienski 1, M C El-Afandi 1, L A Dantuono 1
PMCID: PMC178233  PMID: 8755894

Abstract

A locus encoding polypeptides associated with flagellar structure and function was identified, sequenced, and characterized in Treponema phagedenis and Treponema pallidum. This locus includes homologs of the FlgD, FlgE, MotA, MOB, FliL, and FliM polypeptides found in Salmonella typhimurium and Bacillus subtilis. These polypeptides are extensively conserved between the two treponemes. Several additional polypeptides or unknown function, including Tapl, located upstream of FlgD, and ORF4, located between FlgE and MotA, were also identified. Transcription analysis using RNA PCR indicated that these genes are likely transcribed as part of a single operon and comprise the 5' region of the treponemal fla operon. Primer extension analysis identified a putative promoter, preceding T. phagedenis tap1 in a region of divergent transcription. Pfla resembles the class II or class III motility-related promoters of S. typhimurium. FlgE and Tap1 were further characterized. Western blotting (immunoblotting) indicated that T. pallidum FlgE exhibited an unusual polypeptide ladder that was similar but not identical to that of T. phagedenis. Triton X-114 phase partitioning of T. phagedenis cells coupled with Western blotting revealed that Tap1 was located in the aqueous phase. Computer analysis indicated that Tap1 had no significant membrane spanning regions, suggesting that it resides primarily in the cytoplasm. The organization and expression of this operon are similar in both treponemes but different from those of previously described motility-related operons. These results indicate that despite extensive amino acid sequence conservation, the expression of spirochete flagellar polypeptides is different from that in other bacteria.

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Selected References

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  1. Albertini A. M., Caramori T., Crabb W. D., Scoffone F., Galizzi A. The flaA locus of Bacillus subtilis is part of a large operon coding for flagellar structures, motility functions, and an ATPase-like polypeptide. J Bacteriol. 1991 Jun;173(11):3573–3579. doi: 10.1128/jb.173.11.3573-3579.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Blair D. F., Berg H. C. Mutations in the MotA protein of Escherichia coli reveal domains critical for proton conduction. J Mol Biol. 1991 Oct 20;221(4):1433–1442. doi: 10.1016/0022-2836(91)90943-z. [DOI] [PubMed] [Google Scholar]
  3. Blair D. F., Berg H. C. The MotA protein of E. coli is a proton-conducting component of the flagellar motor. Cell. 1990 Feb 9;60(3):439–449. doi: 10.1016/0092-8674(90)90595-6. [DOI] [PubMed] [Google Scholar]
  4. Canale-Parola E. Motility and chemotaxis of spirochetes. Annu Rev Microbiol. 1978;32:69–99. doi: 10.1146/annurev.mi.32.100178.000441. [DOI] [PubMed] [Google Scholar]
  5. Champion C. I., Miller J. N., Lovett M. A., Blanco D. R. Cloning, sequencing, and expression of two class B endoflagellar genes of Treponema pallidum subsp. pallidum encoding the 34.5- and 31.0-kilodalton proteins. Infect Immun. 1990 Jun;58(6):1697–1704. doi: 10.1128/iai.58.6.1697-1704.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Charon N. W., Goldstein S. F., Block S. M., Curci K., Ruby J. D., Kreiling J. A., Limberger R. J. Morphology and dynamics of protruding spirochete periplasmic flagella. J Bacteriol. 1992 Feb;174(3):832–840. doi: 10.1128/jb.174.3.832-840.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Charon N. W., Goldstein S. F., Curci K., Limberger R. J. The bent-end morphology of Treponema phagedenis is associated with short, left-handed, periplasmic flagella. J Bacteriol. 1991 Aug;173(15):4820–4826. doi: 10.1128/jb.173.15.4820-4826.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Claros M. G., von Heijne G. TopPred II: an improved software for membrane protein structure predictions. Comput Appl Biosci. 1994 Dec;10(6):685–686. doi: 10.1093/bioinformatics/10.6.685. [DOI] [PubMed] [Google Scholar]
  9. Devereux J., Haeberli P., Smithies O. A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):387–395. doi: 10.1093/nar/12.1part1.387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Ge Y., Old I., Saint Girons I., Yelton D. B., Charon N. W. FliH and fliI of Borrelia burgdorferi are similar to flagellar and virulence factor export proteins of other bacteria. Gene. 1996 Feb 2;168(1):73–75. doi: 10.1016/0378-1119(95)00743-1. [DOI] [PubMed] [Google Scholar]
  11. Hardham J. M., Frye J. G., Stamm L. V. Identification and sequences of the Treponema pallidum fliM', fliY, fliP, fliQ, fliR and flhB' genes. Gene. 1995 Dec 1;166(1):57–64. doi: 10.1016/0378-1119(95)00583-x. [DOI] [PubMed] [Google Scholar]
  12. Heinzerling H. F., Penders J. E., Burne R. A. Identification of a fliG homologue in Treponema denticola. Gene. 1995 Aug 8;161(1):69–73. doi: 10.1016/0378-1119(95)00257-7. [DOI] [PubMed] [Google Scholar]
  13. Helmann J. D. Alternative sigma factors and the regulation of flagellar gene expression. Mol Microbiol. 1991 Dec;5(12):2875–2882. doi: 10.1111/j.1365-2958.1991.tb01847.x. [DOI] [PubMed] [Google Scholar]
  14. Helmann J. D., Chamberlin M. J. DNA sequence analysis suggests that expression of flagellar and chemotaxis genes in Escherichia coli and Salmonella typhimurium is controlled by an alternative sigma factor. Proc Natl Acad Sci U S A. 1987 Sep;84(18):6422–6424. doi: 10.1073/pnas.84.18.6422. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Henikoff S. Unidirectional digestion with exonuclease III in DNA sequence analysis. Methods Enzymol. 1987;155:156–165. doi: 10.1016/0076-6879(87)55014-5. [DOI] [PubMed] [Google Scholar]
  16. Homma M., DeRosier D. J., Macnab R. M. Flagellar hook and hook-associated proteins of Salmonella typhimurium and their relationship to other axial components of the flagellum. J Mol Biol. 1990 Jun 20;213(4):819–832. doi: 10.1016/S0022-2836(05)80266-9. [DOI] [PubMed] [Google Scholar]
  17. Homma M., Kutsukake K., Hasebe M., Iino T., Macnab R. M. FlgB, FlgC, FlgF and FlgG. A family of structurally related proteins in the flagellar basal body of Salmonella typhimurium. J Mol Biol. 1990 Jan 20;211(2):465–477. doi: 10.1016/0022-2836(90)90365-S. [DOI] [PubMed] [Google Scholar]
  18. Kihara M., Homma M., Kutsukake K., Macnab R. M. Flagellar switch of Salmonella typhimurium: gene sequences and deduced protein sequences. J Bacteriol. 1989 Jun;171(6):3247–3257. doi: 10.1128/jb.171.6.3247-3257.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Kutsukake K., Doi H. Nucleotide sequence of the flgD gene of Salmonella typhimurium which is essential for flagellar hook formation. Biochim Biophys Acta. 1994 Aug 2;1218(3):443–446. doi: 10.1016/0167-4781(94)90202-x. [DOI] [PubMed] [Google Scholar]
  20. Kutsukake K., Ohya Y., Iino T. Transcriptional analysis of the flagellar regulon of Salmonella typhimurium. J Bacteriol. 1990 Feb;172(2):741–747. doi: 10.1128/jb.172.2.741-747.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Limberger R. J., Charon N. W. Treponema phagedenis has at least two proteins residing together on its periplasmic flagella. J Bacteriol. 1986 Apr;166(1):105–112. doi: 10.1128/jb.166.1.105-112.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Limberger R. J., Slivienski L. L., Samsonoff W. A. Genetic and biochemical analysis of the flagellar hook of Treponema phagedenis. J Bacteriol. 1994 Jun;176(12):3631–3637. doi: 10.1128/jb.176.12.3631-3637.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Limberger R. J., Slivienski L. L., Yelton D. B., Charon N. W. Molecular genetic analysis of a class B periplasmic-flagellum gene of Treponema phagedenis. J Bacteriol. 1992 Oct;174(20):6404–6410. doi: 10.1128/jb.174.20.6404-6410.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Macnab R. M. Genetics and biogenesis of bacterial flagella. Annu Rev Genet. 1992;26:131–158. doi: 10.1146/annurev.ge.26.120192.001023. [DOI] [PubMed] [Google Scholar]
  25. Malakooti J., Komeda Y., Matsumura P. DNA sequence analysis, gene product identification, and localization of flagellar motor components of Escherichia coli. J Bacteriol. 1989 May;171(5):2728–2734. doi: 10.1128/jb.171.5.2728-2734.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Miao R., Fieldsteel A. H. Genetics of Treponema: relationship between Treponema pallidum and five cultivable treponemes. J Bacteriol. 1978 Jan;133(1):101–107. doi: 10.1128/jb.133.1.101-107.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Mirel D. B., Lustre V. M., Chamberlin M. J. An operon of Bacillus subtilis motility genes transcribed by the sigma D form of RNA polymerase. J Bacteriol. 1992 Jul;174(13):4197–4204. doi: 10.1128/jb.174.13.4197-4204.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Norris S. J., Charon N. W., Cook R. G., Fuentes M. D., Limberger R. J. Antigenic relatedness and N-terminal sequence homology define two classes of periplasmic flagellar proteins of Treponema pallidum subsp. pallidum and Treponema phagedenis. J Bacteriol. 1988 Sep;170(9):4072–4082. doi: 10.1128/jb.170.9.4072-4082.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Ohnishi K., Ohto Y., Aizawa S., Macnab R. M., Iino T. FlgD is a scaffolding protein needed for flagellar hook assembly in Salmonella typhimurium. J Bacteriol. 1994 Apr;176(8):2272–2281. doi: 10.1128/jb.176.8.2272-2281.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Oliver D. Protein secretion in Escherichia coli. Annu Rev Microbiol. 1985;39:615–648. doi: 10.1146/annurev.mi.39.100185.003151. [DOI] [PubMed] [Google Scholar]
  31. Pallesen L., Hindersson P. Cloning and sequencing of a Treponema pallidum gene encoding a 31.3-kilodalton endoflagellar subunit (FlaB2). Infect Immun. 1989 Jul;57(7):2166–2172. doi: 10.1128/iai.57.7.2166-2172.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Radolf J. D., Chamberlain N. R., Clausell A., Norgard M. V. Identification and localization of integral membrane proteins of virulent Treponema pallidum subsp. pallidum by phase partitioning with the nonionic detergent triton X-114. Infect Immun. 1988 Feb;56(2):490–498. doi: 10.1128/iai.56.2.490-498.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Raha M., Sockett H., Macnab R. M. Characterization of the fliL gene in the flagellar regulon of Escherichia coli and Salmonella typhimurium. J Bacteriol. 1994 Apr;176(8):2308–2311. doi: 10.1128/jb.176.8.2308-2311.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Sadziene A., Thomas D. D., Bundoc V. G., Holt S. C., Barbour A. G. A flagella-less mutant of Borrelia burgdorferi. Structural, molecular, and in vitro functional characterization. J Clin Invest. 1991 Jul;88(1):82–92. doi: 10.1172/JCI115308. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Stamm L. V., Gherardini F. C., Parrish E. A., Moomaw C. R. Heat shock response of spirochetes. Infect Immun. 1991 Apr;59(4):1572–1575. doi: 10.1128/iai.59.4.1572-1575.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Thomas D. D., Navab M., Haake D. A., Fogelman A. M., Miller J. N., Lovett M. A. Treponema pallidum invades intercellular junctions of endothelial cell monolayers. Proc Natl Acad Sci U S A. 1988 May;85(10):3608–3612. doi: 10.1073/pnas.85.10.3608. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Walker E. M., Arnett J. K., Heath J. D., Norris S. J. Treponema pallidum subsp. pallidum has a single, circular chromosome with a size of approximately 900 kilobase pairs. Infect Immun. 1991 Jul;59(7):2476–2479. doi: 10.1128/iai.59.7.2476-2479.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. di Guan C., Li P., Riggs P. D., Inouye H. Vectors that facilitate the expression and purification of foreign peptides in Escherichia coli by fusion to maltose-binding protein. Gene. 1988 Jul 15;67(1):21–30. doi: 10.1016/0378-1119(88)90004-2. [DOI] [PubMed] [Google Scholar]

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