Purification and some properties of soybean saponin hydrolase from Aspergillus oryzae KO-2.
We had investigated the enzymatic hydrolysis of soybean saponins and selected soybean saponin hydrolase from Aspergillus oryzae KO-2. We attempted purification of this enzyme for further characterization. This enzyme was purified 1500-fold using ammonium sulfate fractionation and Sephadex G-200 gel filtrations. The enzyme was electrophoretically homogeneous and a glycoprotein by PAS staining. By gel filtration, the molecular weight of enzyme was 158,000 and SDS-PAGE showed the enzyme to have a tetrameric structure composed of heterogeneous subunits of 35,000 and 45,000. The enzyme activity was stable at temperatures below 40 degrees C and stable from pH 5.0 to 8.0. The optimum pH was pH 4.5 to 5.0 and the optimum temperature was 50 degrees C. The Km and Vmax for soyasaponin I were 0.48 mM and 9.8 mumol/hr mg protein, respectively. After hydrolysis with the enzyme, soyasapogenol B and alpha-L-rhamnopyranosyl (1----2)-beta-D-galactopyranosyl (1----2)-D-glucuronopyranoside were released from soyasaponin I.
Kudou
S
S
Kanesa Miso Co., Ltd., Aomori, Japan.
Tsuizaki
I
I
Uchida
T
T
Okubo
K
K
D001236:Aspergillus oryzae / Q000201:enzymology*; D002851:Chromatography, High Pressure Liquid; D002855:Chromatography, Thin Layer; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D006867:Hydrolases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000302:isolation & purification*; D007700:Kinetics; D008670:Metals / Q000494:pharmacology; D015394:Molecular Structure; D008970:Molecular Weight; D012503:Saponins / Q000737:chemistry / Q000378:metabolism*; D013025:Glycine max; D013379:Substrate Specificity
D008670:Metals; D012503:Saponins; D006867:Hydrolases