Purification and Properties of Cyclodextrin glycosyltransferase from Bacillus stearothermophilus KY-126

Bacillus stearothermophilus KY-126가 생산하는 Cyclodextrin glycosyltransferase의 정제 및 특성

A bacterial strain No. KY-126, which produced extracellular cyclodextrin glycosyltransferase(CGTase), was isolated from soil and identified as Bacillus stearothermophilus KY-126. The enzyme was purified by the treatments of ammonium sulfate precipitation, DEAF-Sephadex, Sephadex G-100 column chromatography. The optimal pH and temperature for the enzyme activity were pH 5.5 and $65^{\circ}C$, respectively. And the enzyme was stable at pH values from 6.0 to 11.0 at $55^{\circ}C$ for 30 min and stable up to $60^{\circ}C$ for 30 min.. The enzyme was inhibited by $HgCl_{2}$. The molecular weight of the enzyme was estimated to be 67,000 by using SDS-PAGE. The maximum conversion from starch to cyclodextrin (CD) by CGTase was 43% and obtained at 6 hr reaction and the ratio of ${\alpha}-,\;{\beta}-,\;{\gamma}-$, CD production at this time was 2.9 : 2.1 : 1.0.

토양을 대상으로 하여 CGTase를 생산하는 균주를 분리, 선별하여 Bacillus stearothermophilus KY-126을 얻었다. CGTase의 정제는 ammonium sulfate precipitation, ion exchange chromatography, gel filtration의 과정을 통해 분리 정제하여 단일 효소를 얻었으며, 분자량은 약 67,000이었다. 효소 반응의 최적 온도는 $65^{\circ}C$였으며, $55^{\circ}C$에서 30분간 열처리에도 비교적 열에 안정하였다. 최저 활성 pH는 5.5였고 pH5.5에서 10.5까지 비교적 안정하였다. $HgCl_{2}$에 의해 저해를 받았으며, 그 외의 금속 이온에는 저해를 받지 않았다. Soluble starch로부터 CD의 전환율은 43%이었으며, ${\alpha}-:,\;{\beta}-:,\;{\gamma}-$, CD의 생성 비율은 2.9 : 2.1 : 1이었다.