Biological and antigenic characterization of three BApNA-hydrolyzing proteases from the culture supernatant of Porphyromonas gingivalis.
Biological and antigenic distinction of 3-N-alpha-benzoyl-DL-arginine p-nitroanilide (BApNA)-hydrolyzing proteases (Pase-B, Pase-C and Pase-S) isolated from the culture supernatant of Porphyromonas gingivalis were determined. Immunoblotting analysis of these enzymes using a polyclonal antibody against Pase-S, which is a soluble, clostripain-like protease, revealed immunological distinction from Pase-C, a vesicle-associated thiol-protease. Pase-B, a vesicle-associated clostripain-like protease, reacted with the antibody and was also found to contain a considerable amount of carbohydrates in its structure, as compared with the others. Analysis of N-terminal amino acids of Pase-B provided a sequence not found in the SwissProt data bank or previously reported as N-terminal sequences of proteases from P. gingivalis. Pase-S, resembling Pase-B in its hydrolytic specificity, cleaved only arginine residues of peptides and degraded type IV and denatured type I collagen. Pase-C hydrolyzed N-alpha-benzoyl-DL-lysine p-nitroanilide and showed the strongest capacity of degrading native type I collagen. This enzyme was also the only one to possess hemagglutinating activity. Our findings suggest that Pase-S from P. gingivalis is less active than Pase-C and that the enzyme may be an isozyme of Pase-B.
Hinode
D
D
Department of Preventive Dentistry, School of Dentistry, University of Tokushima, Japan.
Masuda
K
K
Yoshioka
M
M
Hayashi
H
H
Nakamura
R
R
Grenier
D
D
Mayrand
D
D
D000345:Affinity Labels; D000595:Amino Acid Sequence; D000942:Antigens, Bacterial / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism*; D001586:Benzoylarginine Nitroanilide / Q000378:metabolism; D015153:Blotting, Western; D050260:Carbohydrate Metabolism; D002241:Carbohydrates / Q000032:analysis; D002364:Caseins / Q000378:metabolism; D003094:Collagen / Q000378:metabolism; D003429:Cross Reactions; D003546:Cysteine Endopeptidases / Q000378:metabolism; D010450:Endopeptidases / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism*; D006384:Hemagglutination; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism; D008239:Lysine / Q000031:analogs & derivatives / Q000378:metabolism; D008969:Molecular Sequence Data; D016966:Porphyromonas gingivalis / Q000201:enzymology* / Q000276:immunology / Q000472:pathogenicity; D011480:Protease Inhibitors / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D000345:Affinity Labels; D000942:Antigens, Bacterial; D001425:Bacterial Outer Membrane Proteins; D002241:Carbohydrates; D002364:Caseins; D007527:Isoenzymes; D011480:Protease Inhibitors; C098918:benzoyllysine nitroanilide; D003094:Collagen; D001586:Benzoylarginine Nitroanilide; D010450:Endopeptidases; C019775:N-benzoyl-DL-arginine-4-nitroanilide hydrolyzing enzyme; D003546:Cysteine Endopeptidases; D008239:Lysine