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PMID: 1322368 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Cloning, expression, and sequencing of a protease gene (tpr) from Porphyromonas gingivalis W83 in Escherichia coli.

Infection and immunity ·Vol. 60 ·No. 8 ·1992-08-00 ·Pages 3186-92

Bourgeau G, Lapointe H, Péloquin P, Mayrand D

Abstract

Porphyromonas gingivalis is a highly proteolytic organism which metabolizes small peptides and amino acids. Indirect evidence suggests that the proteases produced by this microorganism constitute an important virulence factor. In this study, a gene bank of P. gingivalis W83 DNA was constructed by cloning 0.5- to 20-kb HindIII-cut DNA fragments into Escherichia coli DH5 alpha by using the plasmid vector pUC19. A clone expressing a protease from P. gingivalis was isolated on LB agar containing 1% skim milk. The clone contained a 3.0-kb insert that coded for a protease with an apparent molecular mass of 64 kDa. Sequencing part of the 3.0-kb DNA fragment revealed an open reading frame encoding a protein of 482 amino acids with a molecular mass of 62.5 kDa. Putative promoter and termination elements flanking the open reading frame were identified. The activity expressed in E. coli was extensively characterized by using various substrates and protease inhibitors, and the results suggest that it is possibly a thiol protease.

Related Genes
tpr
MeSH Terms
Amino Acid Sequence Base Sequence Cloning, Molecular Endopeptidases/genetics Escherichia coli/genetics Genes, Bacterial Hydrogen-Ion Concentration Molecular Sequence Data Porphyromonas gingivalis/enzymology,genetics Protease Inhibitors/pharmacology Restriction Mapping
Chemicals
Protease Inhibitors Endopeptidases
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Bourgeau G
Département de Biochimie, Faculté des Sciences et de Génie, Université Laval, Québec, Canada.
Lapointe H
Péloquin P
Mayrand D
References (41)
41 references, click to expand
  1. Detection of collagenase activity in oral bacteria.
    Can J Microbiol. 1985 Feb;31(2):134-8 PMID: 2985228
  2. Degradation of immunoglobulins A2, A2, and G by suspected principal periodontal pathogens.
    Infect Immun. 1981 Dec;34(3):757-65 PMID: 7037640
  3. Gene cloning and expression of a Bacteroides gingivalis-specific protein antigen in Escherichia coli.
    Adv Dent Res. 1988 Nov;2(2):310-4 PMID: 3078101
  4. Implantation of Bacteroides gingivalis in nonhuman primates initiates progression of periodontitis.
    Science. 1988 Jan 1;239(4835):55-7 PMID: 3336774
  5. Molecular cloning of two cysteine proteinases from paw-paw (Carica papaya).
    Biochem J. 1986 Jul 1;237(1):105-10 PMID: 3541893
  6. DNA sequencing with chain-terminating inhibitors.
    Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463-7 PMID: 271968
  7. Subgingival microflora and periodontal disease.
    J Clin Periodontol. 1979 Oct;6(5):351-82 PMID: 393729
  8. Isolation, expression, and nucleotide sequence of the sod gene from Porphyromonas gingivalis.
    Infect Immun. 1991 Apr;59(4):1564-6 PMID: 1840572
  9. Cloning and sequencing of the class B beta-lactamase gene (ccrA) from Bacteroides fragilis TAL3636.
    Antimicrob Agents Chemother. 1990 Aug;34(8):1590-2 PMID: 2121094
  10. Proteases and their inhibitors in chronic inflammatory periodontal disease.
    J Clin Periodontol. 1986 Jan;13(1):19-26 PMID: 2418067
  11. Characterization of sodium dodecyl sulfate-stable Bacteroides gingivalis proteases by polyacrylamide gel electrophoresis.
    Infect Immun. 1989 Jan;57(1):95-9 PMID: 2462541
  12. Possibility of Bacteroides gingivalis hemagglutinin possessing protease activity revealed by inhibition studies.
    Microbiol Immunol. 1989;33(1):75-80 PMID: 2543890
  13. Bacterial adhesion to oral tissues: a model for infectious diseases.
    J Dent Res. 1989 May;68(5):750-60 PMID: 2654229
  14. The expression and function of a Bacteroides gingivalis hemagglutinin gene in Escherichia coli.
    Oral Microbiol Immunol. 1989 Sep;4(3):121-31 PMID: 2700777
  15. Molecular cloning and sequencing of the gene encoding the fimbrial subunit protein of Bacteroides gingivalis.
    J Bacteriol. 1988 Apr;170(4):1658-65 PMID: 2895100
  16. Isolation and characterization of a protease from Bacteroides gingivalis.
    Infect Immun. 1987 Mar;55(3):716-20 PMID: 3546137
  17. Degradation of human immunoglobulins G and M and complement factors C3 and C5 by black-pigmented Bacteroides.
    J Med Microbiol. 1985 Feb;19(1):85-94 PMID: 3968707
  18. Activation of keratinocyte-mediated collagen (type I) breakdown by suspected human periodontopathogen. Evidence of a novel mechanism of connective tissue breakdown.
    J Periodontal Res. 1984 Nov;19(6):645-50 PMID: 6084707
  19. Black-pigmented Bacteroides spp. in the human oral cavity.
    Infect Immun. 1981 Apr;32(1):198-203 PMID: 6111541
  20. Construction and expression of recombinant plasmids encoding type 1 or D-mannose-resistant pili from a urinary tract infection Escherichia coli isolate.
    Infect Immun. 1981 Sep;33(3):933-8 PMID: 6116675
  21. Degradation in vivo of the C3 protein of guinea-pig complement by a pathogenic strain of Bacteroides gingivalis.
    Scand J Dent Res. 1984 Feb;92(1):14-24 PMID: 6372060
  22. A comprehensive set of sequence analysis programs for the VAX.
    Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):387-95 PMID: 6546423
  23. Degradation of albumin, haemopexin, haptoglobin and transferrin, by black-pigmented Bacteroides species.
    J Med Microbiol. 1984 Aug;18(1):39-46 PMID: 6748039
  24. Degradation of collagenous substrates by Bacteroides melaninogenicus.
    J Bacteriol. 1961 Apr;81:614-21 PMID: 13705066
  25. Black-pigmented Bacteroides from clinically characterized periodontal sites.
    J Periodontal Res. 1979 Sep;14(5):376-82 PMID: 44523
  26. Structure of actinidin: details of the polypeptide chain conformation and active site from an electron density map at 2-8 A resolution.
    J Mol Biol. 1977 Sep 25;115(3):263-77 PMID: 592367
  27. Hemolytic activity in the periodontopathogen Porphyromonas gingivalis: kinetics of enzyme release and localization.
    Infect Immun. 1991 Jun;59(6):1932-40 PMID: 2037355
  28. Selective modulation of bacterial attachment to oral epithelial cells by enzyme activities associated with poor oral hygiene.
    J Periodontal Res. 1990 May;25(3):172-8 PMID: 2141877
  29. A protease of Bacteroides gingivalis degrades cell surface and matrix glycoproteins of cultured gingival fibroblasts and induces secretion of collagenase and plasminogen activator.
    Infect Immun. 1989 Jan;57(1):213-8 PMID: 2535833
  30. Further studies on the degradation of immunoglobulins by black-pigmented Bacteroides.
    Oral Microbiol Immunol. 1989 Mar;4(1):12-8 PMID: 2628862
  31. Fibronectin fragmentation induced by dental plaque and Bacteroides gingivalis.
    Scand J Dent Res. 1987 Aug;95(4):308-14 PMID: 2820020
  32. The effect of periodontal proteolytic Bacteroides species on proteins of the human complement system.
    J Periodontal Res. 1988 May;23(3):187-92 PMID: 2969970
  33. Identification of cDNA clones encoding a precursor of rat liver cathepsin B.
    Proc Natl Acad Sci U S A. 1985 Apr;82(8):2320-4 PMID: 2986112
  34. Biology of asaccharolytic black-pigmented Bacteroides species.
    Microbiol Rev. 1988 Mar;52(1):134-52 PMID: 3280965
  35. Trypsin-like enzyme activity of the extracellular membrane vesicles of Bacteroides gingivalis W50.
    J Gen Microbiol. 1987 Oct;133(10):2883-94 PMID: 3449601
  36. Selected characteristics of pathogenic and nonpathogenic strains of Bacteroides gingivalis.
    J Clin Microbiol. 1987 Apr;25(4):738-40 PMID: 3571482
  37. The 3'-terminal sequence of Escherichia coli 16S ribosomal RNA: complementarity to nonsense triplets and ribosome binding sites.
    Proc Natl Acad Sci U S A. 1974 Apr;71(4):1342-6 PMID: 4598299
  38. Characterization of a trypsin-like protease from the bacterium Bacteroides gingivalis isolated from human dental plaque.
    Arch Oral Biol. 1984;29(7):559-64 PMID: 6089721
  39. Association of oral Bacteroides with gingivitis and adult periodontitis.
    J Periodontal Res. 1981 May;16(3):259-65 PMID: 6118402
  40. Detection of proteases in polyacrylamide gels containing covalently bound substrates.
    Anal Biochem. 1984 Feb;136(2):470-5 PMID: 6372542
  41. Attachment of Bacteroides gingivalis to collagenous substrata.
    J Dent Res. 1988 Aug;67(8):1075-80 PMID: 3042824
Article Info
Journal
Infection and immunity
Abbr.
Infect Immun
ISSN
0019-9567
DOI
10.1128/iai.60.8.3186-3192.1992
Published
1992-08-00
Pages
3186-92
Language
English
Region
United States
NLM ID
0246127
PMCID
PMC257300
Subset
IM
Databases
GENBANK
M24252, M27037, M27305, M27306, M63610, M63611, M63612, M63613, M63614, M63615, M63616, M63617, M84471, X03970, X63185, Y00641, Y00708
External Links
PubMed source DOI: 10.1128/iai.60.8.3186-3192.1992 PMC Full Text View journal details
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