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Titolo:
TITRATION AND MAPPING OF THE ACTIVE-SITE OF CYSTEINE PROTEINASES FROMPORPHYROMONAS-GINGIVALIS (GINGIPAINS) USING PEPTIDYL CHLOROMETHANES
Autore:
POTEMPA J; PIKE R; TRAVIS J;
Indirizzi:
JAGIELLONIAN UNIV,INST MOL BIOL,DEPT MICROBIOL & IMMUNOL PL-31120 KRAKOW POLAND UNIV CAMBRIDGE,CTR MRC,DEPT HAEMATOL CAMBRIDGE CB2 2QH ENGLAND UNIV GEORGIA,DEPT BIOCHEM & MOL BIOL ATHENS GA 30205
Titolo Testata:
Biological chemistry
fascicolo: 3-4, volume: 378, anno: 1997,
pagine: 223 - 230
SICI:
1431-6730(1997)378:3-4<223:TAMOTA>2.0.ZU;2-B
Fonte:
ISI
Lingua:
ENG
Soggetto:
VASCULAR-PERMEABILITY ENHANCEMENT; BACTEROIDES-GINGIVALIS; ARG-GINGIPAIN; PROTEASE; VIRULENCE; GENE; W83; PURIFICATION; PERIODONTITIS; EXPRESSION;
Keywords:
AFFINITY LABELING; ENZYME INHIBITION; PATHOGENICITY; PERIODONTITIS; PROTEOLYTIC ENZYME; VIRULENCE FACTORS;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Science Citation Index Expanded
Citazioni:
51
Recensione:
Indirizzi per estratti:
Citazione:
J. Potempa et al., "TITRATION AND MAPPING OF THE ACTIVE-SITE OF CYSTEINE PROTEINASES FROMPORPHYROMONAS-GINGIVALIS (GINGIPAINS) USING PEPTIDYL CHLOROMETHANES", Biological chemistry, 378(3-4), 1997, pp. 223-230

Abstract

Porphyromonas gingivalis is one of the major pathogens associated with periodontal disease and releases powerful cysteine proteinases knownas the gingipains, which are key virulence factors for this organism. The three forms of gingipains, gingipain R1, gingipain R2 (gingipain Rs) and gingipain K, which cleave specifically after arginine (R) or lysine (K) residues, were characterized in terms of the kinetics of their interaction with a wide range of synthetic peptidyl chloromethane inhibitors and a peptidyl (acyloxy)-methane. Chloromethane inhibitors were found to inhibit all the enzymes to varying degree dependent on the peptidyl components of the inhibitor. Thus, inhibitors containing a basic residue at P1 rapidly inactivated the gingipains and some specificity could be seen at the P2 site. The (acyloxy)methane inhibitor, Cbz-Phe-Lys-CH2OCO-2,4,6-Me-3-Ph, was very specific in its rapid inhibition of gingipain K over the gingipains R. This inhibitor, together with the peptidyl chloromethanes, D-Phe-Pro-Arg-CHpCI and D-Phe-Phe-Arg-CH2Cl, which reacted most rapidly with the Arg-specific proteinases, could be used to active site titrate purified forms of the enzymes and enzymes found in crude fractions such as intact P. gingivalis cells, vesicles or membrane fractions. From these titrations it was evident that gingipains R were always in an excess of about 3-fold over gingipainK and that the gingipains as a whole made up 85% of the proteolytic activity associated with the bacterium. The elucidation of the kineticsof inhibition by the range of compounds and the development of the titration method for gingipains will considerably aid in future studies on the proteases elaborated by P. gingivalis.

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Documento generato il 27/10/20 alle ore 05:02:07