초록
토양으로부터 levan을 분해하여 단일종의 fructooligosaccharide를 생산하는 새로운 미생물을 분리 선별하여 본 연구의 공시균주로 선택하였다. Levanase 생산을 위한 최적 배지조성은 0.5% levan, 0.3% yeast extract 0.3% $NaNO_3$, 0.1% $K_2HPO_4$, 0.05% NaCl(pH 8.0)이었으며, 500ml용 shaking flask에 배지 50ml를 넣어 $30^{\circ}C$에서 54시간 배양시켰을 때 목적효소의 생산이 최대에 도달하였다. Levanase에 의해 생성되는 생성물은 단일종의 oligo당임이 확인되었으며 측쇄가 많은 levan으로부터는 소량의 측쇄구조를 가진 oligosaccharide로 추정되는 미지의 물질이 부산물로 생성되었다. 생성 oligo당을 순수하게 정제하여 HPLC 및 ESI-MASS로 중합도를 조사한 결과 DP가 7인 levanheptaose임이 판명되었다.
The microorganisms degrading levan were screened from soil. The isolated strain produced levanase releasing single oligosacchride from levan. The optimum cultural medium for levanase production (g/l) was composed of 0.5% levan, 0.1% $K_2HPO_4$, 0.05% NaCl, 0.3% $NaNO_3$, 0.3% yeast extract (pH 8.0). The cultivation for levanase production was carried out in 500 ml shaking flask containing 50 ml of the optimum medium at $30^{\circ}C$ on a reciprocal shaker, and the highest levanase production was observed after 54 hours of cultivation. The levanase hydrolyzed levan into single oligosaccharide. The product purified by chilled EtOH precipitation and gel filtration was detected as a single peak on HPLC analysis. The oligosaccharides formed by enzyme reaction was identified as levanheptaose (DP7) by HPLC and by ESI-MASS.