베타-1,3-글루칸 합성효소 저해제의 스크리닝을 위한 효소원 제조법

Preparation of Enzyme Source for Screening of Enzyme Inhibitor of $\beta$-1,3-glucan Synthase

Assay conditions for screening of $\beta$-1,3-glucan synthase inhibitor were evaluated. Cells in the beginning of mid-log phase showed the highest activity of the $\beta$-1,3-glucan synthase. Cells permeabilized with 1% digitonin treatment could be used as a good crude enzyme source for convenient screening of the $\beta$-1,3-glucan synthase inhibitors. Calcofluor white (0.125% in final) and papulacandin B (25 $\mu$g/ml) inhibit 90% and more than 50% of the $\beta$-1,3-glucan synthase activity, respectively. Cells grown at 37$\circ$C showed higher enzyme activity than those of 25$\circ$C. Catalytic factor of the $\beta$-1,3-glucan synthase was solubilized from particulated membrane preparations, holoenzyme, by extracting with 0.00938% CHAPS.