Immunological characterization and localization of a Porphyromonas gingivalis BApNA-hydrolyzing protease possessing hemagglutinating activity.
A monoclonal antibody (mAb-PC) was produced against a BA pNA-hydrolyzing protease possessing hemagglutinating activity (Pase-C) from Porphyromonas gingivalis. Other P. gingivalis BA pNA-hydrolyzing enzymes (Pase-B and Pase-S) did not react with this antibody. By ELISA or SDS-PAGE and Western immunoblotting analysis, mAb-PC recognized all P. gingivalis and P. endodontalis strains tested but did not recognize other members of the Porphyromonas genus nor other putative periodontopathogenic organisms. Pase-C, extracellular vesicles (ECV) and human strains of P. gingivalis showed two major immunoreactive bands (44 kDa and 40 kDa), whereas a different pattern was obtained with animal strains of P. gingivalis. Biotinylarginyl chloromethane, an irreversible inhibitor of trypsin-like proteases, did not affect the reactivity of Pase-C with mAb-PC on immunoblot. By reversed-phase electronmicroscopy following immunogold labeling, the antibody was shown to bind to the cell surface of P. gingivalis. mAb-PC inhibited the hemagglutinating activity of both P. gingivalis cells and ECV whereas a monoclonal antibody against LPS of P. gingivalis did not. These results suggest that Pase-C is located on the cell surface of P. gingivalis and may participate in erythrocyte binding.
Hinode
D
D
Department of Preventive Dentistry, School of Dentistry, University of Tokushima, Japan.
Masuda
K
K
Yoshioka
M
M
Watanabe
K
K
Umemoto
T
T
Grenier
D
D
Mayrand
D
D
Nakamura
R
R
D000818:Animals; D000911:Antibodies, Monoclonal; D001586:Benzoylarginine Nitroanilide / Q000378:metabolism*; D002415:Cats; D004285:Dogs; D010450:Endopeptidases; D006385:Hemagglutination Inhibition Tests; D006801:Humans; D051379:Mice; D016253:Microscopy, Immunoelectron; D010447:Peptide Hydrolases / Q000276:immunology / Q000378:metabolism*; D017861:Porphyromonas / Q000201:enzymology / Q000302:isolation & purification; D016966:Porphyromonas gingivalis / Q000201:enzymology* / Q000302:isolation & purification / Q000648:ultrastructure; D013045:Species Specificity
D000911:Antibodies, Monoclonal; D001586:Benzoylarginine Nitroanilide; D010450:Endopeptidases; C019775:N-benzoyl-DL-arginine-4-nitroanilide hydrolyzing enzyme; D010447:Peptide Hydrolases