A rapid purification method for soybean Bowman-Birk protease inhibitor using hydrophobic-interaction chromatography.
A protein fraction was isolated from defatted soybean flour by extraction at acid pH, 40% ammonium sulfate precipitation, hydrophobic interaction chromatography, and gel filtration chromatography. SDS-PAGE, under reducing conditions, confirmed it as a homogeneous preparation. This conclusion was consistent with N-terminal amino acid sequence data (20 cycles) which showed a major sequence identical to those reported for soybean Bowman-Birk-type protease inhibitor (BBI), and also indicated a minimum 95% purity based on recoveries of PTH-amino acid residues. The purified fraction inhibited both trypsin and chymotrypsin with average specific activities of 350 and 672 units mg(-1), respectively. Compared with classical BBI purification, this procedure is very rapid requiring only 72-96 h to achieve a yield of 37 mg purified BBI per 200 g starting material.
Yeboah
N A
NA
Genertic Engineering Laboratory, National Food Research Institute, Kannondai, Tsukuba Science City, Ibaraki, Japan.
Arahira
M
M
Udaka
K
K
Fukazawa
C
C
D000595:Amino Acid Sequence; D002364:Caseins / Q000737:chemistry; D002850:Chromatography, Gel; D002918:Chymotrypsin / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D014358:Trypsin Inhibitor, Bowman-Birk Soybean / Q000302:isolation & purification* / Q000378:metabolism
D002364:Caseins; D014358:Trypsin Inhibitor, Bowman-Birk Soybean; D002918:Chymotrypsin