Skip to main content
NCBI home page
Infection and Immunity logoLink to Infection and Immunity
. 1997 Nov;65(11):4888–4891. doi: 10.1128/iai.65.11.4888-4891.1997

Cloning, expression, and sequencing of a protease gene from Bacteroides forsythus ATCC 43037 in Escherichia coli.

T Saito 1, K Ishihara 1, T Kato 1, K Okuda 1
PMCID: PMC175704  PMID: 9353083

Abstract

We have isolated and characterized an N-benzoyl-Val-Gly-Arg-p-nitroanilide-specific protease gene, designated prtH, from Bacteroides forsythus ATCC 43037. Nucleotide sequencing of the DNA insert from the clone (hereafter referred to as clone FST) revealed that the protease activity corresponded to an open reading frame consisting of 1,272 bp coding for a 47.8-kDa protein. When plasmid pFST was used as a probe in Southern hybridization, Sau3AI-digested chromosomal DNA of B. forsythus ATCC 43037 as well as the chromosomal DNAs of the isolated strains Ta4, TR5, and YG2 showed 0.6- and 0.8-kb hybridizing bands. The cell-free extracts of clone FST showed hemolytic activity on human blood cells. The hydrolytic activity of cell extracts of the pFST clone was inhibited by p-toluenesulfonyl-L-lysine chloromethyl ketone hydrochloride, leupeptin, N-ethylmaleimide, iodoacetic acid, iodoaceteamide, and EDTA.

Full Text

The Full Text of this article is available as a PDF (492.9 KB).

Selected References

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

  1. Braun V., Focareta T. Pore-forming bacterial protein hemolysins (cytolysins). Crit Rev Microbiol. 1991;18(2):115–158. doi: 10.3109/10408419109113511. [DOI] [PubMed] [Google Scholar]
  2. Chu L., Bramanti T. E., Ebersole J. L., Holt S. C. Hemolytic activity in the periodontopathogen Porphyromonas gingivalis: kinetics of enzyme release and localization. Infect Immun. 1991 Jun;59(6):1932–1940. doi: 10.1128/iai.59.6.1932-1940.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Dzink J. L., Socransky S. S., Haffajee A. D. The predominant cultivable microbiota of active and inactive lesions of destructive periodontal diseases. J Clin Periodontol. 1988 May;15(5):316–323. doi: 10.1111/j.1600-051x.1988.tb01590.x. [DOI] [PubMed] [Google Scholar]
  4. Grenier D. Characteristics of hemolytic and hemagglutinating activities of Treponema denticola. Oral Microbiol Immunol. 1991 Aug;6(4):246–249. doi: 10.1111/j.1399-302x.1991.tb00485.x. [DOI] [PubMed] [Google Scholar]
  5. Holt S. C., Bramanti T. E. Factors in virulence expression and their role in periodontal disease pathogenesis. Crit Rev Oral Biol Med. 1991;2(2):177–281. doi: 10.1177/10454411910020020301. [DOI] [PubMed] [Google Scholar]
  6. Ishihara K., Kuramitsu H. K. Cloning and expression of a neutral phosphatase gene from Treponema denticola. Infect Immun. 1995 Apr;63(4):1147–1152. doi: 10.1128/iai.63.4.1147-1152.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Ishihara K., Miura T., Kuramitsu H. K., Okuda K. Characterization of the Treponema denticola prtP gene encoding a prolyl-phenylalanine-specific protease (dentilisin). Infect Immun. 1996 Dec;64(12):5178–5186. doi: 10.1128/iai.64.12.5178-5186.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Kadowaki T., Yoneda M., Okamoto K., Maeda K., Yamamoto K. Purification and characterization of a novel arginine-specific cysteine proteinase (argingipain) involved in the pathogenesis of periodontal disease from the culture supernatant of Porphyromonas gingivalis. J Biol Chem. 1994 Aug 19;269(33):21371–21378. [PubMed] [Google Scholar]
  9. Kimizuka R., Miura T., Okuda K. Characterization of Actinobacillus actinomycetemcomitans hemolysin. Microbiol Immunol. 1996;40(10):717–723. doi: 10.1111/j.1348-0421.1996.tb01132.x. [DOI] [PubMed] [Google Scholar]
  10. LISTGARTEN M. A. ELECTRON MICROSCOPIC OBSERVATIONS ON THE BACTERIAL FLORA OF ACUTE NECROTIZING ULCERATIVE GINGIVITIS. J Periodontol. 1965 Jul-Aug;36:328–339. doi: 10.1902/jop.1965.36.4.328. [DOI] [PubMed] [Google Scholar]
  11. Lai C. H., Listgarten M. A., Shirakawa M., Slots J. Bacteroides forsythus in adult gingivitis and periodontitis. Oral Microbiol Immunol. 1987 Dec;2(4):152–157. doi: 10.1111/j.1399-302x.1987.tb00299.x. [DOI] [PubMed] [Google Scholar]
  12. Lisser S., Margalit H. Compilation of E. coli mRNA promoter sequences. Nucleic Acids Res. 1993 Apr 11;21(7):1507–1516. doi: 10.1093/nar/21.7.1507. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Loesche W. J., Syed S. A., Schmidt E., Morrison E. C. Bacterial profiles of subgingival plaques in periodontitis. J Periodontol. 1985 Aug;56(8):447–456. doi: 10.1902/jop.1985.56.8.447. [DOI] [PubMed] [Google Scholar]
  14. Mäkinen P. L., Mäkinen K. K., Syed S. A. An endo-acting proline-specific oligopeptidase from Treponema denticola ATCC 35405: evidence of hydrolysis of human bioactive peptides. Infect Immun. 1994 Nov;62(11):4938–4947. doi: 10.1128/iai.62.11.4938-4947.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Okuda K., Yamamoto A., Naito Y., Takazoe I., Slots J., Genco R. J. Purification and properties of hemagglutinin from culture supernatant of Bacteroides gingivalis. Infect Immun. 1986 Dec;54(3):659–665. doi: 10.1128/iai.54.3.659-665.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Ringquist S., Shinedling S., Barrick D., Green L., Binkley J., Stormo G. D., Gold L. Translation initiation in Escherichia coli: sequences within the ribosome-binding site. Mol Microbiol. 1992 May;6(9):1219–1229. doi: 10.1111/j.1365-2958.1992.tb01561.x. [DOI] [PubMed] [Google Scholar]
  17. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  19. Uitto V. J., Grenier D., Chan E. C., McBride B. C. Isolation of a chymotrypsinlike enzyme from Treponema denticola. Infect Immun. 1988 Oct;56(10):2717–2722. doi: 10.1128/iai.56.10.2717-2722.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Uitto V. J., Pan Y. M., Leung W. K., Larjava H., Ellen R. P., Finlay B. B., McBride B. C. Cytopathic effects of Treponema denticola chymotrypsin-like proteinase on migrating and stratified epithelial cells. Infect Immun. 1995 Sep;63(9):3401–3410. doi: 10.1128/iai.63.9.3401-3410.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Winston J. L., Chen C. K., Neiders M. E., Dyer D. W. Membrane protein expression by Actinobacillus actinomycetemcomitans in response to iron availability. J Dent Res. 1993 Oct;72(10):1366–1373. doi: 10.1177/00220345930720100501. [DOI] [PubMed] [Google Scholar]

Articles from Infection and Immunity are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES