Cyclodextrin분해효소의 정제 및 그 특성

Purification and Some Properties of Cyclodextrin Hydrolase

Bacillus stearothermophilus KFCC 21203를 배양하여 cyclodextrin(CD)을 분해하는 효소를 분리, 정제하고 정제효소의 몇 가지 특성을 조사하였다. 배양액에서 얻은 조효소를 염석, DEAE-cellulose column chromatography, Ultro AcA 34 gel filtration등의 방법으로 15배 정제하였으며 회수율은 77.2%이었다. 정제효소의 specific activity는 12.30units/mg protein 이었고 분자량은 약 29,500정도였다. 이 효소의 작용최적 PH는 5.5, 작용최적온도는 $55^{\circ}C$였으며 $40^{\circ}C$이하의 온도와 pH $5.0{\sim}8.0$의 범위에서 안정하였고 ${\gamma}-CD$에 대한 Km치는 $3.78{\times}10^{-3}$ M이었다. 정제효소는 ${\beta}-CD$에 대한 활성이 매우 낮았고 ${\alpha}-CD$에는 거의 활성이 없었으나 ${\gamma}-CD$에는 매우 높은 활성을 나타내었으며, 이의 분해산물로는 주로 glucose 및 maltose 그리고 소량의 maltotriose였다. 또한 amylose, potato starch, corn starch, amylopectin 및 maltooligomer등에는 높은 활성을, 그리고 glycogen, dextrin에도 비교적 높은 활성을 나타내었고 분해산물로는 주로 glucose와 maltose였다.

Cyclodextrin hydrolase from Bacillus stearothermophilus KFCC 21203 was purified and the properties of the purified enzyme were investigated. The enzyme was purified 15 folds with 77 % recovery by ammonium sulfate fractionation, DEAE-cellulose chromatography, and Ultro AcA 34 gel filtration. The specific activity and the molecular weight of the enzyme were 1.30 units/mg protein and about 29,500, respectively, The maximum activity of the enzyme was shown at $55^{\circ}C$ and pH 5.5. However, stable temperature and pH were $40^{\circ}C$ and $5.0{\sim}8.0$, respectively. The Km value for ${\gamma}-cyclodextrin$ was $3.78{\times}10^{-3}$ M. The degradation activity of the enzyme was selectively high for ${\gamma}-cyclodextrin$, and very low for ${\beta}-cyclodextrin$, but not for ${\alpha}-cyclodextrin$. The decomposed products of ${\gamma}-cyclodextrin$ were mainly glucose and maltose, and a little mlatotriose. The activity of the enzyme was very high for amylose, potato starch, corn starch, amylopectin and maltooligomer, and relatively high for glycogen and dextrin. The decomposed products of them were mainly glucose and maltose.