Purification and characterization of a fibrinogenolytic serine proteinase from Aspergillus fumigatus culture filtrate

FEBS Letters

Bouchara, Jean-Philippe²  Tronchin, Guy²  Chabasse, Dominique²  Annaix, Véronique²  Symoens, Françoise¹  Larcher, Gérald² 

[1] Institute of Hygiene and Epidemiology, 14 rue Juliette Wytsman, B-1050 Bruxelles, Belgium;Laboratoire de Parasitologie-Mycologie, Centre Hospitalier Régional et Universitaire, 4 rue Larrey, F-49033 Angers Cedex, France

关键词: Enzyme purification;    Serine proteinase;    Fibrinogenolytic;    Aspergillus fumigatus;    CBS;    Centraalbureau voor Schimmelcultures;    MCA;    7-amino-4-methyl-coumarin;    PEC;    polyethyleneglycol;    pNA;    paranitroanilide;    SDS-PAGE;    sodium dodecyl sulfate-polyacrylamide gel electrophoresis;   

DOI  :  10.1016/0014-5793(92)81052-N

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

A fibrinogenolytic proteinase has been isolated from Aspergillus fumigatus culture filtrate by ammonium sulfate precipitation followed by successive chromatographics 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°C, although a decrease in activity was observed above 37°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.

【 授权许可】

Unknown