Genetic Analyses of Proteolysis, Hemoglobin Binding, and Hemagglutination of Porphyromonas gingivalis

Shi, Yixin; Ratnayake, Dinath B.; Okamoto, Kuniaki; Abe, Naoko; Yamamoto, Kazuo; Nakayama, Koji

doi:10.1074/jbc.274.25.17955

Cited by 311 publications

(352 citation statements)

References 46 publications

(45 reference statements)

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“…In contrast, characterization of a P. gingivalis rgpA rgpB double mutant has failed to demonstrate a similar role in nutrition for the RGPs either in this study or in an earlier report (Shi et al, 1999). Clearly the possibility that the RGPs play an important nutritional role in vivo through degradation of specific host proteins or release of important protein-bound micronutrients cannot be excluded.…”

Section: Discussioncontrasting

confidence: 49%

“…In addition to a role in haemin acquisition, KGP also appears to be important for growth of P. gingivalis on large proteins since kgp mutants grow very poorly in a defined medium containing albumin as the sole source of carbon and nitrogen ( Fig. 3) and this effect can be reversed by pretreatment of the medium with trypsin (Shi et al, 1999). Hence there is reasonable evidence that KGP has an important nutritional role.…”

Section: Discussionmentioning

confidence: 87%

“…This was confirmed by , who suggested that abnormal processing produced KGP in the RGPnull mutant with reduced activity or rapid degradation of the processed enzyme. Conversely, in a more recent report, Shi et al (1999) observed that an rgpA rgpB mutant of P. gingivalis (KDP133) produced higher levels of KGP activity compared to the parent strain. Similar discrepancies have been reported with respect to Arg-X activity in kgp mutant strains.…”

Section: Introductionmentioning

confidence: 94%

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Generation of Lys-gingipain protease activity in Porphyromonas gingivalis W50 is independent of Arg-gingipain protease activities

et al. 2000

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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.

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Section: Discussioncontrasting

confidence: 49%

Section: Discussionmentioning

confidence: 87%

Section: Introductionmentioning

confidence: 94%

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Generation of Lys-gingipain protease activity in Porphyromonas gingivalis W50 is independent of Arg-gingipain protease activities

et al. 2000

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“…The adhesin domains consist of HGP44, HGP15, HGP17 and HGP27. HGP44 and HGP17 are believed to be involved in haemagglutination, while HGP15 has the ability to bind haemoglobin (Curtis et al, 1996;Kelly et al, 1997;Nakayama et al, 1998;Shi et al, 1999;Shibata et al, 1999).…”

Section: Discussionmentioning

confidence: 99%

Adhesins encoded by the gingipain genes of Porphyromonas gingivalis are responsible for co-aggregation with Prevotella intermedia

et al. 2003

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Co-aggregation among bacterial cells caused by the adherence of one bacterial species to another is a potential colonization mechanism. Several putative aggregation factors for co-aggregation between Porphyromonas (Por.) gingivalis and Prevotella (Pre.) intermedia were partially purified from Por. gingivalis vesicles by gel filtration and affinity chromatography. Antisera against the aggregation factors were made. Analysis using these antisera revealed that 18 and 44 kDa proteins might be responsible for Por. gingivalis vesicle-mediated aggregation of Pre. intermedia. Using antiserum against the 18 kDa protein, the DNA region encoding it was cloned from Por. gingivalis genomic DNA. Sequence analysis revealed that the DNA region was located within the rgpA and kgp genes, encoding Arg-gingipain (Rgp) and Lys-gingipain (Kgp), respectively, and it encoded non-catalytic adhesin domain regions, namely a C-terminal portion of HGP15, the entire HGP17 sequence and an N-terminal portion of HGP27. A portion of the DNA sequence was also found in the haemagglutinin A (hagA) gene. A recombinant glutathione S-transferase (GST)-HGP17 fusion protein reacted to antiserum against the 18 kDa protein and Pre. intermedia cells could adhere to GST-HGP17-conjugated Sepharose 4B beads, indicating that the HGP17 domain protein is responsible for Por. gingivalis vesicle-mediated aggregation of Pre. intermedia.

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“…Periodontopathic bacteria were grown anaerobically at 37°C (80% N 2 , 10% CO 2 , and 10% H 2 ). P. gingivalis ATCC 33277, KDP136 (53), and NDP200 (15) were cultured in enriched brain heart infusion broth (Becton Dickinson, Franklin Lakes, NJ) supplemented with 0.1% cysteine, 5 g/ml of hemin, and 0.5 g/ml of menadione in the absence and presence of appropriate antibiotics (ampicillin, erythromycin, tetracycline, and chloramphenicol) for KDP136 and NDP200, as previously described (15). P. intermedia ATCC 25611 and T. forsythia ATCC 43037, provided by the RIKEN BRC through the National Bio-Resource Project of MEXT, Japan, were cultured in anaerobic bacterial culture media (Eiken Chemical, Tokyo, Japan) supplemented with 1% cysteine and 0.5 g/ml of menadione.…”

Section: Methodsmentioning

confidence: 99%

Degradation of Incretins and Modulation of Blood Glucose Levels by Periodontopathic Bacterial Dipeptidyl Peptidase 4

et al. 2017

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Severe periodontitis is known to aggravate diabetes mellitus, though molecular events related to that link have not been fully elucidated. Porphyromonas gingivalis, a major pathogen of periodontitis, expresses dipeptidyl peptidase 4 (DPP4), which is involved in regulation of blood glucose levels by cleaving incretins in humans. We examined the enzymatic characteristics of DPP4 from P. gingivalis as well as two other periodontopathic bacteria, Tannerella forsythia and Prevotella intermedia, and determined whether it is capable of regulating blood glucose levels. Cellassociated DPP4 activity was found in those microorganisms, which was effectively suppressed by inhibitors of human DPP4, and molecules sized 73 kDa in P. gingivalis, and 71 kDa in T. forsythia and P. intermedia were immunologically detected. The k cat /K m values of recombinant DPP4s ranged from 721 Ϯ 55 to 1,283 Ϯ 23 M Ϫ1 s Ϫ1 toward Gly-Pro-4-methylcoumaryl-7-amide (MCA), while those were much lower for His-Ala-MCA. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) analysis showed His/Tyr-Ala dipeptide release from the N termini of incretins, glucagon-like peptide 1 (GLP-1) and glucose-dependent insulinotropic polypeptide, respectively, with the action of microbial DPP4. Moreover, intravenous injection of DPP4 into mice decreased plasma active GLP-1 and insulin levels, accompanied by a substantial elevation in blood glucose over the control after oral glucose administration. These results are the first to show that periodontopathic bacterial DPP4 is capable of modulating blood glucose levels the same as mammalian DPP4; thus, the incidence of periodontopathic bacteremia may exacerbate diabetes mellitus via molecular events of bacterial DPP4 activities.

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Genetic Analyses of Proteolysis, Hemoglobin Binding, and Hemagglutination of Porphyromonas gingivalis

Cited by 311 publications

References 46 publications

Generation of Lys-gingipain protease activity in Porphyromonas gingivalis W50 is independent of Arg-gingipain protease activities

Generation of Lys-gingipain protease activity in Porphyromonas gingivalis W50 is independent of Arg-gingipain protease activities

Adhesins encoded by the gingipain genes of Porphyromonas gingivalis are responsible for co-aggregation with Prevotella intermedia

Degradation of Incretins and Modulation of Blood Glucose Levels by Periodontopathic Bacterial Dipeptidyl Peptidase 4

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