미생물에 의한 $\beta$-Galactosidase의 생산 및 이용에 관한 연구 (제 1보) Penicillium sp.로부터 효소의 생산조건 및 정제

Studies on the Production of $\beta$-Galactosidase by Microorganism and its Application (Part 1) Conditions for the Production and Purification of the Enzyme from Penicillium SP.

토양에서 분리된 여러가지 곰팡이류 가운데 $\beta$-galactosidase 분비력이 강하고 중성부근에서 효소안정성이 높은 Penicillium sp. 균주를 $\beta$-galactosidase 생산균주로 선정하였다. 이 균주는 밀기울고체배지에서 3$0^{\circ}C$ 72시간 배양에서 galactosidase 분비력이 가장 높았으며 질소원 첨가배지에서 질소원의 종류에 따라 14%~85%까지 증가되었다. 고체배지에서 얻은 효소추출액을 ammonium sulfate fractionation, SP-Sephadex C-50 chromatography, ultrogel AcA44 filtration, hrdroxrapatite chromatography등에 의하여 비활성도는 101 units/mg protein으로 5050배 정제되었다. 최종적으로 정제된 galactosidase는 analytical ultracentrifuge에서 단일단백으로 Schlieren pattern을 나타냈고 Disc electrophoresis에서는 단일 band로서 전기영동되었으며 영동된 $\beta$-galactosidase 활성 band와 일치하였다.

A strain of Penicillium sp. which produces considerable amount of $\beta$-galactosidase was selected from extracellular $\beta$-galactosidase-producing fungi isolated from soil. The enzyme was found to be very stable in neutral pH range. Maximum enzymatic activity was reached after 72 hr of incubation in a wheat bran medium at 3$0^{\circ}C$. Productivity of the enzyme appeared not to be affected by the addition of carbon sources to the medium but the activity of the enzyme was increased from 14% to 85% by the addition of various nitrogen sources. The enzyme extracted from the wheat-bran culture of the Penicillium sp. was purified to 5050-fold by ammonium sulfate fractionation, SP-Sephadex C-50 chromatography, Ultrogel AcA 44 filtration and hydroxyapatite chromatography. The purified $\beta$-galactosidase was homogeneous on ultracentrifugation and disc electrophoresis.