A consensus Porphyromonas gingivalis promoter sequence.

We have determined the transcription start points (tsp) for recently identified Porphyromonas gingivalis W50 genes, kgp, rgpA, rgpB (formerly designated prtK, prtR, and prtRII respectively), fetB and the mcmAB operon. Alignment of the DNA upstream of these tsp and those from the literature has enabled us to identify consensus sequences that may represent a P. gingivalis promoter. There is a potential -10 hexamer sequence, 5'-TATATT-3' centred on average at -10/11 nt which is repeated at -19/20 nt and an upstream consensus, 5'-CAGAT(A/G)-3' which is centred at -39/40 nt.

Serum immunoglobulin G (IgG) and IgG subclass responses to the RgpA-Kgp proteinase-adhesin complex of Porphyromonas gingivalis in adult periodontitis.

Serum immunoglobulin G (IgG), IgM, and IgG subclass responses to the RgpA-Kgp proteinase-adhesin complex of Porphyromonas gingivalis were examined by enzyme-linked immunosorbent assay using adult periodontitis patients and age- and sex-matched controls. Twenty-five sera from subjects with adult periodontitis (diseased group) and 25 sera from healthy subjects (control group) were used for the study. Sera and subgingival plaque samples from 10 sites were collected from each patient at the time of clinical examination. The level of P. gingivalis in the plaque samples was determined using a DNA probe. Highly significant positive associations between the percentage of sites positive for P. gingivalis and measures of disease severity (mean pocket depth, mean attachment loss, and percentage of sites that bled on probing) were found. The diseased group had significantly higher specific IgG responses to the RgpA-Kgp complex than did the control group, and the responses were significantly associated with mean probing depths and percentage of sites positive for P. gingivalis. Analysis of the IgG subclass responses to the RgpA-Kgp complex revealed that the subclass distribution for both the diseased and control groups was IgG4 > IgG2 > IgG3 = IgG1. The IgG2 response to the complex was positively correlated with mean probing depth, whereas the IgG4 response was negatively correlated with this measure of disease severity. Immunoblot analysis of the RgpA-Kgp complex showed that sera from healthy subjects and those with low levels of disease, with high IgG4 and low IgG2 responses, reacted with the RgpA27, Kgp39, and RgpA44 adhesins; however, sera from diseased subjects with low IgG4 and high IgG2 responses reacted only with the RgpA44 and/or Kgp44 adhesins. Epitope mapping of the RgpA27 adhesin localized a major epitope recognized by IgG4 antibodies in sera from subjects with high IgG4 and low IgG2 responses to the RgpA-Kgp complex which was not recognized by sera from diseased subjects with low IgG4 and high IgG2 responses.

Characterization of a Porphyromonas gingivalis gene prtK that encodes a lysine-specific cysteine proteinase and three sequence-related adhesins.

Porphyromonas gingivalis extracellular arginine- and lysine-specific proteinases have been implicated as major virulence factors in the development of adult periodontitis. We have previously purified a 48-kDa lysine-specific cysteine proteinase, designated PrtK48, from a P. gingivalis W50 cell-associated multiprotein complex. PrtK48 was non-covalently associated with three sequence-related adhesins designated PrtK39, PrtK15 and PrtK44 in the multiprotein complex. In this study we cloned and characterized the gene, designated prtK, that encodes a polyprotein that is post-translationally processed to yield the Lys-specific proteinase PrtK48 and the three sequence-related adhesins PrtK39, PrtK15 and PrtK44.

Characterization of a Porphyromonas gingivalis gene prtR that encodes an arginine-specific thiol proteinase and multiple adhesins.

Cysteine proteinases of Porphyromonas gingivalis have been implicated as major virulence factors in the development of periodontitis. Several groups have reported the characterisation of similar genes encoding the same arginine-specific thiol proteinase from P. gingivalis; however, the reported size and structure of the genes have varied. We report here the complete nucleotide sequence of the gene prtR that encodes a polyprotein containing the Arg-specific proteinase and multiple haemagglutinins/adhesins. The nascent polyprotein consists of a putative leader sequence and a prosequence followed by the 45 kDa Arg-specific proteinase and 44, 15, 17 and 27 kDa sequence-related adhesins in that order. The size and structure of the prtR are consistent with the size of the mRNA transcript (5.3 kb) and the size and sequences of the individual protein components purified from P. gingivalis.

Complete nucleotide sequence of a gene prtR of Porphyromonas gingivalis W50 encoding a 132 kDa protein that contains an arginine-specific thiol endopeptidase domain and a haemagglutinin domain.

We have purified from Porphyromonas gingivalis W50 a 45 kDa arginine-specific, thiol-activated, EDTA-sensitive endopeptidase, designated prtR. Oligonucleotide probes based on the N-terminal amino acid sequence were used to isolate a genomic fragment containing an open reading frame (3654 bp) with the potential to encode a 132 kDa protein including the prtR N-terminus. Analysis of this prtR gene revealed that the predicted nascent product contains a protease domain followed by a haemagglutinin domain and is post-translationally processed by proteolytic (possibly autolytic) events to produce a 43-54 kDa arginine-specific, thiol protease and a 41-53 kDa haemagglutinin. Comparison of the prtR with the P. gingivalis prtH gene suggests that the prtH gene product also contains protease and haemagglutinin domains but in the reverse order to that in the prtR. An overlapping but shifted reading frame at the 3' end of the prtR encodes the 5' region of the prtH.