A STUDY ON α-AMYLASE FROM BACILLUS SUBTILIS 86315 II.PURIFICATION,PROPERTIES AND KINETICS

α-Amylase from Bacillus subtilis 86315 was purified by means of chitosan flocculation, starch adsorption, ethanol precipitation, Sephadex G-100 gel filtration, and DEAE-cellulose chromatography. The purified α-amylase showed a single band on polyacrylamide gel electrophoresis (PAGE). The molecular weight of this α-amylase was estimated to be about 54 000 by SDS-PAGE. The pI of the enzyme was 5.2 by isoelectric focusing. The enzyme has an optimum temperature of 60℃ and is stable below 50℃. Its optimum pH is 5.6 and the enzyme is stable from pH 5.6 to 8.8. Km value of this enzyme for soluble starch as a substrate is 3.0mg/ml. Ca~(2+), Ba~(2+), Mg~(2+) stimulate the α-amylase activity slightly. Mn~(2+) has no effect on the activity of this α-amylase. Fe~(2+), Fe~(3+), Cu~(2+), EDTA, formaldehyde and glutaraldehyde strongly inhibit the α-amylase activity. The enzyme has a high degree of stability in 60％ ethanol whereas it can be denatured casily by urea.