Purification and characterization of a strong fibrinolytic enzyme (nattokinase) in the vegetable cheese natto, a popular soybean fermented food in Japan.
A strong fibrinolytic enzyme (nattokinase) was purified from the vegetable cheese natto. Nattokinase was extracted from natto with saline and isolated by sequential use of hydrophobic chromatography on Butyl-Toyopearl, ion-exchange chromatography on CM-Toyopearl, and gel-filtration on Sephadex G-50. The isolated protein gave a single sharp band on SDS-PAGE either before or after reduction. The sequence, as determined by automated Edman degradation of the uncleaved molecule and its enzymatically derived peptide, consisted of a total 275 amino acid residues (M.W = 27,728) and exhibited a high homology with the subtilisins. The purified nattokinase digested not only fibrin but also several synthetic substrates. Among the synthetic substrates, the most sensitive substrate was Suc-Ala-Ala-Pro-Phe-pNA for subtilisin. PMSF inhibited both the fibrinolytic activity and the amidolytic activity. The results indicate that nattokinase is a subtilisin-like serine protease.
Fujita
M
M
Biotechnology Research Laboratories, JCR Pharmaceuticals Co., Ltd., Kobe, Japan.
Nomura
K
K
Hong
K
K
Ito
Y
Y
Asada
A
A
Nishimuro
S
S
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D005337:Fibrin / Q000378:metabolism; D005342:Fibrinolysis; D005343:Fibrinolytic Agents / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D005511:Food Handling; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009842:Oligopeptides / Q000378:metabolism; D012697:Serine Endopeptidases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D013025:Glycine max / Q000201:enzymology*; D013379:Substrate Specificity; D013381:Subtilisins
D005343:Fibrinolytic Agents; D009842:Oligopeptides; D005337:Fibrin; D012697:Serine Endopeptidases; D013381:Subtilisins; C053771:nattokinase