糸状菌の生産するセルラーゼに関する研究 : (第3報)ペニシリウムセルラーゼの分離精製とその性質について

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As reported previously, the cellulase system in a preparation from Penicillium variabile is capable of digesting native cellulose completely into glucose, and is composed mainly of three components. In this report, are described the results of further studies on the fractionation and properties of the components.The fraction I isolated by means of adsorption on cellulose powder in M/100 acetate buffer at pH 5.0 followed by elution with Tween 20 (500 ppm) is rich in C_1-enzyme which is further purified by gel filtration. It is capable of dissolving cellulose powder giving rise to oligosaccharides together with a very small amount of glucose, but has a very low activity in digesting CMC as measured by reducing sugar formation and viscosity lowering. The enzyme has a Mol. Wt. of ca. 25,000 as estimated by the gel filtration technique with Sephadex.The fraction II, which is much less adsorbable on cellulose powder than the fraction I, is adsorbed effectively on phosphoric acid-swollen cellulose in M/100 acetate buffer at pH 5.0 and is eluted therefrom by water. It is rich in C_x-enzyme which is further purified by gel filtration. The enzyme attacks modified cellulose; viscosity lowering and reducing sugar formation are remarkable with CMC as substrate. It hydrolyzes cellopen taose, cellotetraose and cellotriose, but is quite inactive in hydrolyzing native cellulose, cellobiose, p-nitrophenyl-β-glucoside (PNPG) and p-nitrophenyl-β-cellobioside (PNPC). By means of gel filtration, the fraction II can be fractionated further into three subfractions having Mol. Wt. of ca. 40,000,60,000 and 80,000,respectively. The first and the third subgractions are major. No appreciable difference is observed in substrate specificity, opt. pH and Km-value (with CMC) among these major subfractions except for thermostability. Also no evidence is obtained for the transformation of the above-mentioned C_x-isozymes from one to another despite a number of tests for the action of protease, PCMB, urea and heat. The effluent pattern for the subfractions changes with the age of culture.The fraction III, which is not absorbed on swollen cellulose as well as cellulose powder, is rich in cellobiase which is further purified by gel filtration. The enzyme is capable of hydrolyzing cello-oligosaccharides such as cellobiose, cellotriose, cellotetraose, cellopentaose, p-nitrophenyl-β-glucoside and p-nitrophenyl-β-cellobioside.The action of the above C_1-enzyme on native cellulose is accelerated by C_x-enzyme but not by cellobiase. With CMC as substrate, cellobiase is highly effective in accelerating the aciton of C_x; e. g. the reaction velocity is raised four-fold. It is obvious therefore that there are conspicuous synergisms between C_1 and C_x for oligosaccharide formation and between C_x and cellobiase for glucose formation.
社団法人日本生物工学会の論文
1967-09-25

糸状菌の生産するセルラーゼに関する研究 : (第3報)ペニシリウムセルラーゼの分離精製とその性質について

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