The prpR1 and prR2 arginine‐specific protease genes of Porphyromonas gingivalis W50 produce five biochemically distinct enzymes

Rangarajan, Minnie; Aduse-Opoku, Joseph; Slaney, J. M.; Young, Kung Chia; Curtis, Michael A.

doi:10.1046/j.1365-2958.1997.2831647.x

Cited by 76 publications

(111 citation statements)

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“…Both of these enzymes are products of two distinct but related genes (27). rgpA encodes a polyprotein which, after post-translational processing/modifications, yields three different forms of the enzyme (28,29); the major one is a non-covalent complex containing separate catalytic and adhesion/hemagglutinin domains (HRgpA). In contrast, the fragment encoding the latter domain(s) is missing in the rgpB gene structure, and its transThis work was supported by Grant 11670219 from the Japanese Ministry of Education (to T.…”

Section: S534 -S536) Our Results Indicate That Bacterial Proteimentioning

confidence: 99%

Activation of Human Prothrombin by Arginine-specific Cysteine Proteinases (Gingipains R) from Porphyromonas gingivalis *

Imamura

Bańbuła

Pereira

et al. 2001

Journal of Biological Chemistry

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The effect of 95-(HRgpA) and 50-kDa gingipain R (RgpB), arginine-specific cysteine proteinases from periodontopathogenic bacterium Porphyromonas gingivalis on human prothrombin activation was investigated. Each enzyme released thrombin from prothrombin in a dose-and time-dependent manner with the former enzyme, containing adhesion domains, being 17-fold more efficient than the single chain RgpB. A close correlation between the generation of fibrinogen clotting activity and amidolytic activity indicated that ␣-thrombin was produced by gingipains R, and this was confirmed by SDS-polyacrylamide gel electrophoresis, thrombin active site labeling, and amino-terminal sequence analysis of prothrombin digestion fragments. Significantly, the catalytic efficiency of HRgpA to generate thrombin (k cat /K m ‫؍‬ 1.2 ؋ 10 6 M ؊1 s ؊1 ) was 100-fold higher than that of RgpB (k cat /K m ‫؍‬ 1.2 ؋ 10 4 M ؊1 s ؊1 ). The superior prothrombinase activity of HRgpA over RgpB correlates with the fact that only the former enzyme was able to clot plasma, and kinetic data indicate that prothrombin activation can occur in vivo. At P. gingivalis-infected periodontitis sites HRgpA may be involved in the direct production of thrombin and, therefore, in the generation of prostaglandins and interleukin-1, both have been found to be associated with the development and progression of the disease. Furthermore, by taking into account that the P. gingivalis bacterium has been immunolocalized in carotid atherosclerotic plaques at thrombus formation sites (Chiu, B. (1999) Am. Heart J. 138, S534 -S536), our results indicate that bacterial proteinases may potentially participate in the pathogenesis of cardiovascular disease associated with periodontitis.

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Section: S534 -S536) Our Results Indicate That Bacterial Proteimentioning

confidence: 99%

Activation of Human Prothrombin by Arginine-specific Cysteine Proteinases (Gingipains R) from Porphyromonas gingivalis *

Imamura

Bańbuła

Pereira

et al. 2001

Journal of Biological Chemistry

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“…These large fragments have never been observed in preparations of RI from P. gingiualis W50. In all previous work (Rangarajan et al, 1997b;Pavloff et al, 1995), the largest peptide from the p-and y-regions of PrpRI has been the p-adhesin domain of RI from P. gingiualis W50.…”

Section: Genetic Origin Of Purified Proteinsmentioning

confidence: 99%

“….) is located in the pro-region of prR2 [Rangarajan et al, 1997b;accession (N. Slakeski and others, complete cds)] and would be generated by processing at Arg,,-Gly,, and Arg,,,-Tyr231, which would release a peptide fragment of 201 residues (Fig. 4).…”

Section: Genetic Origin Of Purified Proteinsmentioning

confidence: 99%

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

1998

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Proteases of Porphyromonas gingivalis are considered to be important factors in the virulence of this organism. A non-pigmenting mutant of P. gingivaris W50 (W50/BE1) has been shown to be less virulent in animal models and to produce significantly less Arg-specif ic protease activity than the parent strain. Three proteases are present in the culture supernatant of P. gingivak W50: RI, RIA and RIB. All three proteases are derived from prpR1, which encodes a polypeptide of 1706 amino acids that is organized into distinct domains (pro, a, B and 7). The aim of the present investigation was to purify and characterize the Arg-specif ic proteases produced b y the avirulent W50/BE1 strain. Significant differences were observed between the proteases of P. gingivalis 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, prR2, and was present in two fractions, RllAs/BE (soluble) and RIIAv/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 PrRll 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 prpR1 enzymes, which is only partially compensated by synthesis of prR2-derived enzymes, and that all of these proteases undergo altered post-translational modification compared to the parent strain.

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“…Furthermore, we define the immunodominant epitope utilizing truncation and alanine scanning techniques and evaluate the epitope immunogenicity in a peptide polymer vaccine construct. (32) and Kgp Ϫ (K1A) (33) were grown as previously described (34). P. gingivalis RgpA-Kgp, RgpA, and Kgp were prepared and characterized as previously described (34).…”

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confidence: 99%

Characterization of T Cell Responses to the RgpA-Kgp Proteinase-Adhesin Complexes of Porphyromonas gingivalis in BALB/c Mice

Tam

O'Brien-Simpson

Pathirana

et al. 2008

The Journal of Immunology

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Porphyromonas gingivalis is a Gram-negative bacterium strongly associated with chronic periodontitis, an inflammatory oral disease. A major virulence factor common to all characterized strains of P. gingivalis is the RgpA-Kgp proteinase-adhesin complexes (RgpA-Kgp complexes). In this study, we investigated T cell proliferative and cytokine responses to the RgpA-Kgp complexes and identified T cell epitopes in BALB/c mice utilizing Pepscan methodology. T cell proliferative responses were found to be predominantly directed toward the proteinase catalytic domains. Eleven T cell epitopes were identified using RgpA-Kgp-primed lymph node T cells (IL-4 dominant) and 21 using an RgpA-Kgp-specific T cell line (IFN-γ dominant), with 5 T cell epitopes, including the immunodominant epitope peptide 22, common to both T cell populations. Peptide 22 (439ANYTAHGSETAWADP453) from the Kgp proteinase catalytic domain induced a Th2 cytokine response in mice, and peptide 22-primed T cells had a Th2 cytokine profile when stimulated with the RgpA-Kgp complexes. Truncation and alanine scanning of peptide 22 identified the minimum epitope (442TAHGSETAWA451), and residues His444, Glu447, and Trp450 as critical for T cell proliferation. With a view to vaccine development, peptide 22 was incorporated into a synthetic peptide polymer. Peptide 22 polymer induced strong T cell proliferation and crossreactivity to native RgpA-Kgp complexes. In conclusion, we have identified a major T cell epitope of P. gingivalis and established that antigenicity of the T cell epitope is retained when delivered as a peptide polymer. The strategies employed here may have potential in the development of a synthetic peptide vaccine for P. gingivalis.

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The prpR1 and prR2 arginine‐specific protease genes of Porphyromonas gingivalis W50 produce five biochemically distinct enzymes

Cited by 76 publications

References 30 publications

Activation of Human Prothrombin by Arginine-specific Cysteine Proteinases (Gingipains R) from Porphyromonas gingivalis *

Activation of Human Prothrombin by Arginine-specific Cysteine Proteinases (Gingipains R) from Porphyromonas gingivalis *

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

Characterization of T Cell Responses to the RgpA-Kgp Proteinase-Adhesin Complexes of Porphyromonas gingivalis in BALB/c Mice

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