Purification and characterization of fibrinolytic alkaline protease from Fusarium sp. BLB

• Published in: Applied microbiology and biotechnology 2007-02, Vol.74 (2), p.331-338
• Main Authors: Ueda, Mitsuhiro, Kubo, Toshihiro, Miyatake, Kazutaka, Nakamura, Takumi
• Format: Article
• Language: English
• Subjects: Alkaline serine protease; Amino Acid Sequence; Amino acids; Bacterial Proteins - antagonists & inhibitors; Bacterial Proteins - chemistry; Bacterial Proteins - isolation & purification; Bacterial Proteins - metabolism; Biological and medical sciences; Biotechnology; Bos taurus; Endopeptidases - chemistry; Endopeptidases - isolation & purification; Endopeptidases - metabolism; Enzyme Stability; Enzymes; Fibrin - metabolism; Fibrinolytic activity; Filtrate; Fundamental and applied biological sciences. Psychology; Fusarium; Fusarium - classification; Fusarium - enzymology; Fusarium - genetics; Fusarium - isolation & purification; Hibiscus; Hibiscus - microbiology; Hydrogen-Ion Concentration; Lumbricus rubellus; Molecular Sequence Data; Molecular weight; Pelamis; Plant Leaves - microbiology; Protease Inhibitors - pharmacology; Proteins; Sequence Analysis, DNA; Serine Endopeptidases - chemistry; Serine Endopeptidases - drug effects; Serine Endopeptidases - isolation & purification; Serine Endopeptidases - metabolism; Streptomyces griseus
• ISSN: 0175-7598; 1432-0614
• DOI: 10.1007/s00253-006-0621-1

Fusarium sp. BLB, which produces a strongly fibrinolytic enzyme, was isolated from plant leaf (Hibiscus). Fibrinolytic alkaline protease was purified from a culture filtrate of Fusarium sp. BLB by precipitation with (NH₄)₂SO₄ and column chromatography with CM-Toyopearl 650M and Superdex 75. The purified enzyme was homogeneous on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The molecular weight was 27,000 by SDS-PAGE. Maximum activity of protease was observed at pH 9.5 and 50°C. Purified protease was active between pH 2.5 and 11.5 and was found to be stable up to 50°C. The enzyme derived from Fusarium sp. BLB is useful for thrombolytic therapy because this enzyme showed pH resistance. The activity was inhibited by diisopropylfluorophosphate and phenylmethylsulfonyl fluoride. The N-terminal amino acid sequence of the enzyme showed a similarity to those of proteases from Fusarium sp., Streptomyces griseus, Bos taurus bovine, Katsuwo pelamis digestive tract, and Lumbricus rubellus.