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Volume 144, Issue 9

Altered expression and modification of proteases from an avirulent mutant of Porphyromonas gingivalis W50 (W50/BE1) Free

Abstract

Proteases of are considered to be important factors in the virulence of this organism. A non-pigmenting mutant of W50 (W50/BE1) has been shown to be less virulent in animal models and to produce significantly less Arg-specific protease activity than the parent strain. Three proteases are present in the culture supernatant of W50: RI, RIA and RIB. All three proteases are derived from which encodes a polypeptide of 1706 amino acids that is organized into distinct domains (pro, α, β and γ). The aim of the present investigation was to purify and characterize the Arg-specific proteases produced by the avirulent W50/BE1 strain. Significant differences were observed between the proteases of W50 and W50/BE1. The levels of RI present in the culture supernatant of W50/BE1 were lower than those present in W50, and RIA and RIB were absent. RI from W50/BE1 was composed of three polypeptide chains, unlike the enzyme from W50, which is a heterodimer. The remainder of the Arg-specific protease activity in W50/BE1 was derived from a second gene, and was present in two fractions, RIIA/BE (soluble) and RIIA/BE (vesicle-bound). This activity contained two peptide chains: a ~ 54 kDa chain corresponding to the protease domain and a ~ 26 kDa chain, derived from the propeptide domain of the PrRII precursor. No enzyme with large glycan additions, equivalent to RIB in the vesicle fraction of the wild-type W50, was present. These data indicate that the reduced level of extracellular protease activity in W50/BE1 reflects reduced synthesis and/or export of enzymes, which is only partially compensated by synthesis of -derived enzymes, and that all of these proteases undergo altered post-translational modification compared to the parent strain.

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Keyword(s): altered modification , avirulent , extracellular arginine-specific proteases and Porphyromonas gingivalis
© Society for General Microbiology 1998
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1998-09-01
2024-04-18
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References

  1. Aduse-Opoku J., Muir J., Slaney J.M., Rangarajan M., Curtis M. 1995; Characterisation, genetic analysis and expression of a protease antigen (PrpRl) of Porphyromonas gingivalis W50.. Infect Immun 63:4744–4754
     [Google Scholar]
  2. Aduse-Opoku J., Rangarajan M., Young K.A., Curtis M.A. 1998; Maturation of the arginine-specific proteases of Porphyromonas gingivalis W50 is dependent on a functional prR2 protease gene.. Infect Immun 66:1594–1600
     [Google Scholar]
  3. Chandad F., Mouton C. 1995; Antigenic, structural, and functional relationships between fimbriae and the hemag- glutinating adhesin HA-Ag2 of Porphyromonas gingivalis.. Infect Immun 63:4755–4763
     [Google Scholar]
  4. Curtis M.A., Aduse-Opoku J., Slaney J.M., Rangarajan M., Booth V., Cridland J., Shepherd P. 1996; Characterization of an adherence and antigenic determinant of the ArgI protease of Porphyromonas gingivalis which is present on multiple gene- products.. Infect Immun 64:2532–2539
     [Google Scholar]
  5. Cutler C.W., Eke P.I., Genco C.A., VanDyke T.E., Arnold R.R. 1996; Hemin-induced modifications of the antigenicity and hemin-binding capacity of Porphyromonas gingivalis lipopoly- saccharide.. Infect Immun 64:2282–2287
     [Google Scholar]
  6. Dawson R.M.C., Elliot D.C., Elliot W.H., Jones K.M. (editors) 1986 Data for Biochemical Research, 3rd edn.. Oxford: Clarendon Press;
     [Google Scholar]
  7. Fletcher H.M., Schenkein H.A., Morgan R.M., Bailey K.A., Berry C.R., Macrina F.L. 1995; Virulence of a Porphyromonas gingivalis W83 mutant defective in the prtH gene.. Infect Immun 63:1521–1528
     [Google Scholar]
  8. Fox T., de Miguel E., Mort J.S., Storer A.C. 1992; Potent slow-binding inhibition of cathepsin B by its propeptide.. Biochemistry 31:12571–12576
     [Google Scholar]
  9. Good N.E., Winget G.D., Winter W., Connolly T.N., Izawa S., Singh R.M.M. 1966; Hydrogen ion buffers for biological research.. Biochemistry 5:467–477
     [Google Scholar]
  10. Han N., Whitlock J., Progulske-Fox A. 1996; The hemag-glutinin gene A (hagA) of Porphyromonas gingivalis 381 contains four large, contiguous, direct repeats.. Infect Immun 64:4000–4007
     [Google Scholar]
  11. Hinode D., Masuda K., Yoshioka M., Watanabe K., Umemoto T., Grenier D., Mayrand D., Nakamura R. 1995; Immunological characterization and localization of a Porphyromonas gingivalis BApNA-hydrolyzing protease possessing hemag- glutinating activity.. FEMS Microbiol Lett 131:211–217
     [Google Scholar]
  12. Imamura T., Potempa J., Tanase S., Travis J. 1997; Activation of blood coagulation factor X by arginine-specific cysteine proteinases (gingipain-R’s) from Porphyromonas gingivalis.. J Biol Chem 27:16062–16067
     [Google Scholar]
  13. Kesavalu L., Holt S.C., Ebersole J.L. 1996; Trypsin-likeprotease activity of Porphyromonas gingivalis as a potential virulence factor in a murine lesion model.. Microh Pathog 20:1–10
     [Google Scholar]
  14. Laemmli U.K. 1970; Cleavage of structural proteins during the assembly of the head of bacteriophage T4.. Nature 227:680–685
     [Google Scholar]
  15. Lindsay S.S., Wheeler B., Sanderson K.E., Costerson J.W. 1973; The release of alkaline phosphatase and of lipopoly- saccharide during the growth of rough and smooth strains of Salmonella typhimurium.. Can J Microbiol 19:335–343
     [Google Scholar]
  16. McKee A.S., McDermid A.S., Wait R., Baskerville A., Marsh P.D. 1988; Isolation of colonial variants of Bacteroides gingivalis W50 with reduced virulence.. J Med Microbiol 27:59–64
     [Google Scholar]
  17. Nakayama K., Kadowaki T., Okamoto K., Yamamoto K. 1995; Construction and characterization of arginine-specific cysteine proteinase(Arg-gingipain)-deficient mutants ofPorphyromonas gingivalis.. J Biol Chem 270:23619–23626
     [Google Scholar]
  18. Nakayama K., Yoshimura F., Kadowaki T., Yamamoto K. 1996; Involvement of arginine-specific cysteine proteinase (Arg- gingipain) in fimbriation of Porphyromonas gingivalis.. J Bacteriol 178:2818–2824
     [Google Scholar]
  19. Ohta Y., Inouye M. 1990; Pro-subtilisin E: purification and characterization of its autoprocessing to active subtilisin E in vitro.. Mol Microbiol 4:295–304
     [Google Scholar]
  20. Okamoto K., Kadowaki T., Nakayama K., Yamamoto K. 1996; Cloning and sequencing of the gene encoding a novel lysine-specific cysteine proteinase (Lys-gingipain) in Porphyromonas gingivalis: structural relationship with the arginine- specific cysteine proteinase (Arg-gingipain).. J Biochem 120:398–406
     [Google Scholar]
  21. Pavloff N., Potempa J., Pike R.N., Prochazka V., Kiefer M.C, Travis J., Barr P.J. 1995; Molecular cloning and structural characterization of the Arg-gingipain proteinase of Porphyromonas gingivalis.. J Biol Chem 270:1007–1010
     [Google Scholar]
  22. Pike R., McGraw W., Potempa J., Travis J. 1994; Lysine- and arginine-specific proteinases from Porphyromonas gingivalis.. J Biol Chem 269:406–411
     [Google Scholar]
  23. Pike R.N., Potempa J., McGraw W., Coetzer T.H.T., Travis J. 1996; Characterization of the binding activities of proteinase- adhesin complexes from Porphyromonas gingivalis.. J Bacteriol 178:2876–2882
     [Google Scholar]
  24. Potempa J., Pike R., Travis J. 1997; Titration and mapping of the active site of cysteine proteinases from Porphyromonas gingivalis (gingipains) using peptidyl chloromethanes.. J Biol Chem 378:223–230
     [Google Scholar]
  25. Rangarajan M., Smith S.J.M, U S., Curtis M.A. 1997a; Biochemical characterization of the arginine-specific proteases of Porphyromonas gingivalis W50 suggests a common precursor.. Biochem J 323:701–709
     [Google Scholar]
  26. Rangarajan M., Aduse-Opoku J., Slaney J.M., Young K.A., Curtis M.A. 1997b; The prpRl and prR2 arginine-specific proteases of Porphyromonas gingivalis W50 produce five biochemically distinct enzymes.. Mol Microbiol 23:955–965
     [Google Scholar]
  27. Shah H.N., Seddon S.V., Gharbia S.E. 1989; Studies on the virulence properties and metabolism of pleiotropic mutants of Porphyromonas gingivalis (Bacteroides gingivalis) W50.. Oral Microbiol Immunol 4:19–23
     [Google Scholar]
  28. Smalley J.W., Birss A.J. 1991; Extracellular vesicle-associated and soluble trypsin-like enzyme fractions of Porphyromonas gingivalis W50.. Oral Microbiol Immunol 6:202–208
     [Google Scholar]
  29. Smalley J.W., Birss A.J., Kay H.M., McKee A.S., Marsh P.D. 1989; The distribution of trypsin-like enzyme activity in cultures of a virulent and an avirulent strain of Bacteroides gingivalis W50.. Oral Microbiol Immunol 4:178–181
     [Google Scholar]
  30. van Steenbergen T.J.N., Kastelein P., Touw J.J.A., de Graff J. 1982; Virulence of black-pigmented Bacteroides strains from periodontal pockets and other sites in experimentally induced skin lesions in mice.. J Periodontal Res 17:41–49
     [Google Scholar]
  31. Zhu X., Ohta Y., Jordan F., Inouye M. 1989; Pro-sequence of subtilisin can guide the refolding of denatured subtilisin in an intramolecular process.. Nature 339:483–484
     [Google Scholar]
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Altered expression and modification of proteases from an avirulent mutant of Porphyromonas gingivalis W50 (W50/BE1)
Microbiology 144, 2487 (1998); https://doi.org/10.1099/00221287-144-9-2487
/content/journal/micro/10.1099/00221287-144-9-2487
/content/journal/micro/10.1099/00221287-144-9-2487
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