Skip to main content
NCBI home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1997 Oct;179(19):6092–6099. doi: 10.1128/jb.179.19.6092-6099.1997

The FliO, FliP, FliQ, and FliR proteins of Salmonella typhimurium: putative components for flagellar assembly.

K Ohnishi 1, F Fan 1, G J Schoenhals 1, M Kihara 1, R M Macnab 1
PMCID: PMC179513  PMID: 9324257

Abstract

The flagellar genes fliO, fliP, fliQ, and fliR of Salmonella typhimurium are contiguous within the fliLMNOPQR operon. They are needed for flagellation but do not encode any known structural or regulatory components. They may be involved in flagellar protein export, which proceeds by a type III export pathway. The genes have been cloned and sequenced. The sequences predict proteins with molecular masses of 13,068, 26,755, 9,592, and 28,933 Da, respectively. All four gene products were identified experimentally; consistent with their high hydrophobic residue content, they segregated with the membrane fraction. From N-terminal amino acid sequence analysis, we conclude that fliO starts immediately after fliN rather than at a previously proposed site downstream. FliP existed in two forms, a 25-kDa form and a 23-kDa form. N-terminal amino acid analysis of the 23-kDa form demonstrated that it had undergone cleavage of a signal peptide--a rare process for prokaryotic cytoplasmic membrane proteins. Site-directed mutation at the cleavage site resulted in impaired processing, which reduced, but did not eliminate, complementation of a fliP mutant in swarm plate assays. A cloned fragment encoding the mature form of the protein could also complement the fliP mutant but did so even more poorly. Finally, when the first transmembrane span of MotA (a cytoplasmic membrane protein that does not undergo signal peptide cleavage) was fused to the mature form of FliP, the fusion protein complemented very weakly. Higher levels of synthesis of the mutant proteins greatly improved function. We conclude that, for insertion of FliP into the membrane, cleavage is important kinetically but not absolutely required.

Full Text

The Full Text of this article is available as a PDF (1.5 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Broome-Smith J. K., Gnaneshan S., Hunt L. A., Mehraein-Ghomi F., Hashemzadeh-Bonehi L., Tadayyon M., Hennessey E. S. Cleavable signal peptides are rarely found in bacterial cytoplasmic membrane proteins (review). Mol Membr Biol. 1994 Jan-Mar;11(1):3–8. doi: 10.3109/09687689409161023. [DOI] [PubMed] [Google Scholar]
  2. Carpenter P. B., Zuberi A. R., Ordal G. W. Bacillus subtilis flagellar proteins FliP, FliQ, FliR and FlhB are related to Shigella flexneri virulence factors. Gene. 1993 Dec 31;137(2):243–245. doi: 10.1016/0378-1119(93)90014-t. [DOI] [PubMed] [Google Scholar]
  3. Engelman D. M., Steitz T. A., Goldman A. Identifying nonpolar transbilayer helices in amino acid sequences of membrane proteins. Annu Rev Biophys Biophys Chem. 1986;15:321–353. doi: 10.1146/annurev.bb.15.060186.001541. [DOI] [PubMed] [Google Scholar]
  4. Fan F., Macnab R. M. Enzymatic characterization of FliI. An ATPase involved in flagellar assembly in Salmonella typhimurium. J Biol Chem. 1996 Dec 13;271(50):31981–31988. doi: 10.1074/jbc.271.50.31981. [DOI] [PubMed] [Google Scholar]
  5. Fickett J. W. Recognition of protein coding regions in DNA sequences. Nucleic Acids Res. 1982 Sep 11;10(17):5303–5318. doi: 10.1093/nar/10.17.5303. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Galán J. E. Molecular genetic bases of Salmonella entry into host cells. Mol Microbiol. 1996 Apr;20(2):263–271. doi: 10.1111/j.1365-2958.1996.tb02615.x. [DOI] [PubMed] [Google Scholar]
  7. Homma M., Iino T., Macnab R. M. Identification and characterization of the products of six region III flagellar genes (flaAII.3 through flaQII) of Salmonella typhimurium. J Bacteriol. 1988 May;170(5):2221–2228. doi: 10.1128/jb.170.5.2221-2228.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Homma M., Kutsukake K., Iino T. Structural genes for flagellar hook-associated proteins in Salmonella typhimurium. J Bacteriol. 1985 Aug;163(2):464–471. doi: 10.1128/jb.163.2.464-471.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Jones C. J., Homma M., Macnab R. M. L-, P-, and M-ring proteins of the flagellar basal body of Salmonella typhimurium: gene sequences and deduced protein sequences. J Bacteriol. 1989 Jul;171(7):3890–3900. doi: 10.1128/jb.171.7.3890-3900.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Jones C. J., Macnab R. M., Okino H., Aizawa S. Stoichiometric analysis of the flagellar hook-(basal-body) complex of Salmonella typhimurium. J Mol Biol. 1990 Mar 20;212(2):377–387. doi: 10.1016/0022-2836(90)90132-6. [DOI] [PubMed] [Google Scholar]
  11. Kawagishi I., Homma M., Williams A. W., Macnab R. M. Characterization of the flagellar hook length control protein fliK of Salmonella typhimurium and Escherichia coli. J Bacteriol. 1996 May;178(10):2954–2959. doi: 10.1128/jb.178.10.2954-2959.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Kawagishi I., Müller V., Williams A. W., Irikura V. M., Macnab R. M. Subdivision of flagellar region III of the Escherichia coli and Salmonella typhimurium chromosomes and identification of two additional flagellar genes. J Gen Microbiol. 1992 Jun;138(6):1051–1065. doi: 10.1099/00221287-138-6-1051. [DOI] [PubMed] [Google Scholar]
  13. 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]
  14. Kutsukake K., Ohya Y., Yamaguchi S., Iino T. Operon structure of flagellar genes in Salmonella typhimurium. Mol Gen Genet. 1988 Sep;214(1):11–15. doi: 10.1007/BF00340172. [DOI] [PubMed] [Google Scholar]
  15. Macnab R. M. The end of the line in bacterial sensing: the flagellar motor. Cold Spring Harb Symp Quant Biol. 1988;53(Pt 1):67–75. doi: 10.1101/sqb.1988.053.01.011. [DOI] [PubMed] [Google Scholar]
  16. Malakooti J., Ely B., Matsumura P. Molecular characterization, nucleotide sequence, and expression of the fliO, fliP, fliQ, and fliR genes of Escherichia coli. J Bacteriol. 1994 Jan;176(1):189–197. doi: 10.1128/jb.176.1.189-197.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Moore K. E., Miura S. A small hydrophobic domain anchors leader peptidase to the cytoplasmic membrane of Escherichia coli. J Biol Chem. 1987 Jun 25;262(18):8806–8813. [PubMed] [Google Scholar]
  18. Müller V., Jones C. J., Kawagishi I., Aizawa S., Macnab R. M. Characterization of the fliE genes of Escherichia coli and Salmonella typhimurium and identification of the FliE protein as a component of the flagellar hook-basal body complex. J Bacteriol. 1992 Apr;174(7):2298–2304. doi: 10.1128/jb.174.7.2298-2304.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Pugsley A. P. The complete general secretory pathway in gram-negative bacteria. Microbiol Rev. 1993 Mar;57(1):50–108. doi: 10.1128/mr.57.1.50-108.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Raha M., Kihara M., Kawagishi I., Macnab R. M. Organization of the Escherichia coli and Salmonella typhimurium chromosomes between flagellar regions IIIa and IIIb, including a large non-coding region. J Gen Microbiol. 1993 Jul;139(7):1401–1407. doi: 10.1099/00221287-139-7-1401. [DOI] [PubMed] [Google Scholar]
  21. 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]
  22. Salmond G. P., Reeves P. J. Membrane traffic wardens and protein secretion in gram-negative bacteria. Trends Biochem Sci. 1993 Jan;18(1):7–12. doi: 10.1016/0968-0004(93)90080-7. [DOI] [PubMed] [Google Scholar]
  23. Steinrücke P., Steffens G. C., Panskus G., Buse G., Ludwig B. Subunit II of cytochrome c oxidase from Paracoccus denitrificans. DNA sequence, gene expression and the protein. Eur J Biochem. 1987 Sep 15;167(3):431–439. doi: 10.1111/j.1432-1033.1987.tb13356.x. [DOI] [PubMed] [Google Scholar]
  24. Stout V., Torres-Cabassa A., Maurizi M. R., Gutnick D., Gottesman S. RcsA, an unstable positive regulator of capsular polysaccharide synthesis. J Bacteriol. 1991 Mar;173(5):1738–1747. doi: 10.1128/jb.173.5.1738-1747.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Studier F. W., Moffatt B. A. Use of bacteriophage T7 RNA polymerase to direct selective high-level expression of cloned genes. J Mol Biol. 1986 May 5;189(1):113–130. doi: 10.1016/0022-2836(86)90385-2. [DOI] [PubMed] [Google Scholar]
  26. Sugimoto K., Sugisaki H., Okamoto T., Takanami M. Studies on bacteriophage fd DNA. IV. The sequence of messenger RNA for the major coat protein gene. J Mol Biol. 1977 Apr 25;111(4):487–507. doi: 10.1016/s0022-2836(77)80065-x. [DOI] [PubMed] [Google Scholar]
  27. Suzuki T., Komeda Y. Incomplete flagellar structures in Escherichia coli mutants. J Bacteriol. 1981 Feb;145(2):1036–1041. doi: 10.1128/jb.145.2.1036-1041.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Van Gijsegem F., Gough C., Zischek C., Niqueux E., Arlat M., Genin S., Barberis P., German S., Castello P., Boucher C. The hrp gene locus of Pseudomonas solanacearum, which controls the production of a type III secretion system, encodes eight proteins related to components of the bacterial flagellar biogenesis complex. Mol Microbiol. 1995 Mar;15(6):1095–1114. doi: 10.1111/j.1365-2958.1995.tb02284.x. [DOI] [PubMed] [Google Scholar]
  29. Vogler A. P., Homma M., Irikura V. M., Macnab R. M. Salmonella typhimurium mutants defective in flagellar filament regrowth and sequence similarity of FliI to F0F1, vacuolar, and archaebacterial ATPase subunits. J Bacteriol. 1991 Jun;173(11):3564–3572. doi: 10.1128/jb.173.11.3564-3572.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Wilson M. L., Macnab R. M. Overproduction of the MotA protein of Escherichia coli and estimation of its wild-type level. J Bacteriol. 1988 Feb;170(2):588–597. doi: 10.1128/jb.170.2.588-597.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Yamaguchi S., Fujita H., Sugata K., Taira T., Iino T. Genetic analysis of H2, the structural gene for phase-2 flagellin in Salmonella. J Gen Microbiol. 1984 Feb;130(2):255–265. doi: 10.1099/00221287-130-2-255. [DOI] [PubMed] [Google Scholar]
  32. von Heijne G. Control of topology and mode of assembly of a polytopic membrane protein by positively charged residues. Nature. 1989 Oct 5;341(6241):456–458. doi: 10.1038/341456a0. [DOI] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES