title: Purification and characterization of a fibrinogenolytic serine proteinase from Aspergillus fumigatus culture filtrate.
abstract: A fibrinogenolytic proteinase has been isolated from Aspergillus fumigatus culture filtrate by ammonium sulfate precipitation followed by successive chromatographies on Sephadex G-75 and immobilized phenylalanine. The purified proteinase exhibited a molecular weight of about 33 kDa. When analysed by SDS-polyacrylamide gels containing co-polymerized fibrinogen, the proteinase appeared as a broad band at the top of the gels, which could correspond to polymerization of the enzyme, as suggested by SDS-PAGE analysis of the unboiled eluate. The isoelectric point was 8.75 and the enzyme was not glycosylated. Proteinase activity was optimum at pH 9 and between 37 and 42 degrees C, although a decrease in activity was observed above 37 degrees C. PMSF and chymostatin markedly inhibited the proteinase activity, and good kinetic constants were obtained for the synthetic substrate, N-Suc-Ala-Ala-Pro-Phe-pNA. These results provide direct evidence that this enzyme belongs to the chymotrypsin-like serine proteinase group.
authors.affiliation: Laboratoire de Parasitologie-Mycologie, Centre Hospitalier Régional et Universitaire, Angers, France.
authors.forename: G
authors.initials: G
authors.lastname: Larcher
authors.forename: J P
authors.initials: JP
authors.lastname: Bouchara
authors.forename: V
authors.initials: V
authors.lastname: Annaix
authors.forename: F
authors.initials: F
authors.lastname: Symoens
authors.forename: D
authors.initials: D
authors.lastname: Chabasse
authors.forename: G
authors.initials: G
authors.lastname: Tronchin
mesh_terms: D000595:Amino Acid Sequence; D001232:Aspergillus fumigatus / Q000201:enzymology*; D002853:Chromatography, Liquid; D003470:Culture Media; D004591:Electrophoresis, Polyacrylamide Gel; D005340:Fibrinogen / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D007700:Kinetics; D008969:Molecular Sequence Data; D012697:Serine Endopeptidases / Q000302:isolation & purification* / Q000378:metabolism; D013696:Temperature
chemical_list: D003470:Culture Media; D005340:Fibrinogen; D012697:Serine Endopeptidases