Porphyromonas gingivalis, a black-pigmenting anaerobe implicated in the aetiology of periodontal disease, contains two loci, rgpA and rgpB, encoding the extracellular Arg-X specific proteases (RGPs, Arg-gingipains), and kgp, which encodes a Lys-X specific protease (KGP, Lys-gingipain). The rgpA and kgp genes encode polyproteins comprising pro-peptide and catalytic domain with large N- and C-terminal extensions which require proteolytic processing at several Arg and Lys residues to generate mature enzymes. The product of rgpB contains only a pro-peptide and the catalytic domain which requires processing at an Arg residue to generate active enzyme. An rgpA rgpB double mutant (E8) of P. gingivalis was constructed to study the role of RGPs in the processing of KGP. A kgp mutant (K1A) was also studied to investigate the role of KGP in the generation of RGPs. E8 was stable in the absence of the antibiotics tetracycline and clindamycin (selection markers for rgpA and rgpB, respectively) and exhibited the same pigmentation, colony morphology and identical growth rates to the parent W50 strain in the absence of antibiotics, in both complex and chemically defined media. The KGP activity of E8, grown in the absence of tetracycline, in whole cultures and in culture supernatants (up to 6 d) was identical to levels in W50. However, in the presence of tetracycline in the growth medium, the level of KGP was reduced to 50% of levels present in whole cultures of W50. Since tetracycline had no effect on RGP or KGP activity when incorporated into assay buffer, this effect is most likely to be on the synthesis of Kgp polypeptide. K1A was also stable in the absence of antibiotics but was unable to pigment, and remained straw-coloured throughout growth. RGP activity in whole cultures of K1A was identical to levels in W50, but RGP activity in 6 d culture supernatants was reduced to 50% of levels present in W50. Thus, although KGP is not required for generation of RGP activity from RgpA and RgpB polypeptides, its absence affects the release/transport of RGP into culture supernatant.
Aduse-OpokuJ., MuirJ., SlaneyJ. M., RangarajanM., CurtisM. A.1995; Characterization, genetic analysis, and expression of a protease antigen (PrpRI) of Porphyromonas gingivalis W50. Infect Immun 63:4744–4754
Aduse-OpokuJ., RangarajanM., YoungK. A., CurtisM. A.1998; Maturation of the arginine-specific proteases of Porphyromonas gingivalis W50 is dependent on functional prR2 protease gene. Infect Immun 66:1594–1600
Birkedal-HansenH., TaylorR. E., ZambonJ. J., BarwaP. K., NeidersM. E.1988; Characterization of collagenolytic activity from strains of Bacteroides gingivalis. J Periodontal Res 23:258–264[CrossRef]
CarlssonJ., HermannB. F., HoflingJ. F., SundqvistG. K.1984; Degradation of the human proteinase inhibitors alpha-1-antitrypsin and alpha-2-macroglobulin by Bacteroides gingivalis. Infect Immun 43:644–648
CurtisM. A., KuramitsuH., LantzM., MacrinaF., NakayamaK., PotempaJ., ReynoldsE., Aduse-OpokuJ.1999; Molecular genetics and nomenclature of proteases of Porphyromonas gingivalis. J Periodontal Res 34:464–472[CrossRef]
DzinkJ. L., SocranskyS. S., HaffajeeA. D.1988; The predominant cultivable microbiota of active and inactive lesions of destructive periodontal-diseases. J Clin Periodontol 15:316–323[CrossRef]
KadowakiT., NakayamaK., YoshimuraF., OkamotoK., AbeN., YamamotoK.1998; Arg-gingipain acts as a major processing enzyme for various cell surface proteins in Porphyromonas gingivalis. J Biol Chem 273:29072–29076[CrossRef]
KatoJ., TakahashiN., KuramitsuH.1992; Sequence analysis and characterization of the Porphyromonas gingivalis prtC gene which expresses a novel collagenase activity. J Bacteriol 174:3889–3895
MaleyJ., ShoemakerN. B., RobertsI. S.1992; Introduction of colonic-Bacteroides shuttle plasmids into Porphyromonas gingivalis: identification of a putative P. gingivalis insertion-sequence element. FEMS Microbiol Lett 93:219–224
MilnerP., BattenJ. E., CurtisM. A.1996; Development of a simple chemically defined medium for Porphyromonas gingivalis: requirement for α-ketoglutarate. FEMS Microbiol Lett 140:125–130
NakayamaK.1997; Domain-specific rearrangement between the two Arg-gingipain-encoding genes in Porphyromonas gingivalis: possible involvement of nonreciprocal recombination. Microbiol Immunol 41:185–196[CrossRef]
NakayamaK., RatnayakeD. B., TsukubaT., KadowakiT., YamamotoK., FujimuraS.1998; Haemoglobin receptor protein is intragenically encoded by the cysteine proteinase-encoding genes and the haemagglutinin-encoding gene of Porphyromonas gingivalis. Mol Microbiol 27:51–61[CrossRef]
OkamotoK., KadowakiT., NakayamaK., YamamotoK.1996; Cloning and sequencing of the gene encoding a novel lysine-specific cysteine protease (Lys-gingipain) in Porphyromonas gingivalis: structural relationship with the arginine-specific cysteine protease (Arg-gingipain). J Biochem 120:398–406[CrossRef]
PavloffN., PembertonP. A., PotempaJ., ChenW.-C. A., PikeR. N., ProchazkaV., KieferM. C., TravisJ., BarrP. J.1997; Molecular cloning and characterization of Porphyromonas gingivalis Lys-gingipain. A new member of an emerging family of pathogenic bacterial cysteine proteinases. J Biol Chem 272:1595–1600[CrossRef]
PikeR., McGrawW., PotempaJ., TravisJ.1994; Lysine- and arginine-specific proteinases from Porphyromonas gingivalis. Isolation, characterization and evidence for the existence of complexes with hemagglutinins. J Biol Chem 269:406–411
RangarajanM., SmithS. J. M., US., CurtisM. A.1997a; Biochemical characterization of the arginine-specific proteases of Porphyromonas gingivalis W50 suggests a common precursor. Biochem J 323:701–709
RangarajanM., Aduse-OpokuJ., SlaneyJ. M., YoungK. A., CurtisM. A.1997b; The prpR1 and prR2 arginine-specific protease genes of Porphyromonas gingivalis W50 produce five biochemically distinct enzymes. Mol Microbiol 23:955–966[CrossRef]
SatoM., OtsukaM., MaeharaR., EndoJ., NakamuraR.1987; Degradation of human secretory immunoglobulin A by protease isolated from the anaerobic periodontopathogenic bacterium, Bacteroides gingivalis. Arch Oral Biol 32:235–238[CrossRef]
SchenkeinH. A., BerryC. R.1988; Production of chemotactic factors for neutrophils following the interaction of Bacteroides gingivalis with purified C5. J Periodontal Res 23:187–192[CrossRef]
ShiY., RatnayakeD. B., OkamotoK., AbeN., YamamotoK., NakayamaK.1999; Genetic analysis of proteolysis, hemoglobin binding, and hemagglutination of Porphyromonasgingivalis. Construction of mutants with a combination of rgpA, rgpB, kgp and hagA. J Biol Chem 274:17955–17960[CrossRef]
SlakeskiN., ClealS. M., BhogalP. S., ReynoldsE. C.1999; Characterisation of a Porphyromonas gingivalis gene prtK that encodes a lysine specific cysteine proteinase and three sequence-related adhesins. Oral Microbiol Immunol 14:92–97[CrossRef]
SlotsJ., BragdL., WikströmM., DahlenG.1986; The occurrence of Actinobacillus actinomycetemcomitans, Bacteroides gingivalis and Bacteroides intermedius in destructive periodontal disease in adults. J Clin Periodontol 13:576–577
SmalleyJ. W., BirssA. J., ShuttleworthC. A.1988; The degradation of type-1 collagen and human plasma fibronectin by the trypsin-like enzyme and extracellular membrane vesicles of Bacteroides gingivalis W50. Arch Oral Biol 33:323–329[CrossRef]
SundqvistG., CarlssonJ., HermannB., TarnvikA.1985; Degradation of human immunoglobulins G and M and complement factors C3 and C5 by black-pigmented Bacteroides. J Med Microbiol 19:85–94[CrossRef]
UittoV.-J., LarjavaH., HeinoJ., SorsaT.1989; 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 57:213–218
WingroveJ. A., DiScipioR. G., ChenZ., PotempaJ., TravisJ., HugliT. E.1992; Activation of complement components C3 and C5 by a cysteine proteinase (gingipain-1) from Porphyromonas (Bacteroides) gingivalis. J Biol Chem 267:18902–18907