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A component of the microcrystalline cellulose (Avicel)-hydrolyzing enzyme produced by a fungal strain, Y-94, tentatively called Avicelase II, was purified by several kinds of column chromatography followed by chromatofocusing withPolybuffer anion exchanger (PBE 94), BioGel AO.5 m gel filtration, electrophoresis on polyacrylamide gel and disc isoelectric focusing (IEF). The· molecular weight of the enzyme was estimated to be 68,000 by the SDS-polyacrylamide gel method and 56,000 by Bio-Gel AO.5 m gel filtration, and its isoelectric point was found to be around 4.4 by the chromatofocusing and disc IEF methods. The optimum pH and temperature for Avicel hydrolysis were 5.3 and 62°C, respectively. The enzyme was stable between pH 4.1 and 6.0 at 4°C for 24hr, and up to 61°C for 10min. Heavy metal ions such as Cu 2 + and Hg 2 + strongly inactivated the enzyme. A highly purified enzyme preparation showed hydrolyzing activities towards Avicel, xylan, carboxylmethyl cellulose (CMC) (activity ratio of 100: 50: 20, respectively) and acidswollen cellulose. By the PAS method, staining glycoprotein with Shiff's reagent and periodic acid, the enzyme seemed to be a glycoprotein containing some carbohydrate in its structure. Avicelase II tended to show a higher affinity towards cellooligosaccharides of high molecular weight. The enzyme hydrolyzed Avicel and acid-swollen cellulose, and produced trace amounts of cellotriose and significant quantities of glucose. In addition, the enzyme is able to act on the internal glycosidic bonds of CMC and xylan. Judging from the results, it appears that Avicelase II should be classified as a member of the specific endoglucanases rather than as an exoglucanase. 2493

Cyclomaltodextrin glucanotransferase was purified from B. circulans C31 through two successive steps of starch and Biogel column chromatography. The enzyme was purified up to 90fold with a 30% yield. Its molecular weight was around 103,000. The purified enzyme converted 28% of the soluble starch to /?-cyclodextrin at pH 7.0 and a substrate concentration of 5%. The optimum pH for the enzymewas found to be 5.5. The optimum temperature was 60°C. The enzyme was stable from pH 5.5~9.0 and up to 50°C. Cyclomaltodextrin glucanotransferase (EC 2.4.1.19, CGTase)*1 catalyzes the formation of cyclodextrins from starch and related carbohydrates such as amylose, amylopectin, glycogen and maltooligosaccharides.1* Certain species of Bacillus have been found to produce CGTases which may differ from one another in various properties and especially in the ratio of cyclodextrin products, a : /? : y formed.2~15) Although studies on CGTases from B. macerans and an alkalophilic Bacillus (ATCC 21783) have been by very extensive,2~4'10~13) investigations of those from other strains have not been reported much. As to B. circulans, only one study on CGTase was reported, by Okada and Kitahata.5) In our previous study,16) a Bacills species was selected for its high-CGTase producting ability. It closely resembled B. circulans as to

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A mesophilic fungal strain Y-94 produced three types of thermostable endo-xylanases accompanied by the formation of a large amount of cellulase. These xylanases were separated from the cellulase by heat treatment at 65°C for 2.5 hr and purified by DEAE-Toyopearl chromatography, chromatography on an anion exchanger (PBE 94), and Bio-Gel A 0.5 m gel filtration. The molecular weights of the three types of xylanase, designated as xylanase A, B and C, were 51,000, 48,000, and 35,000, respectively. All three enzymes showed highest activity at pH 4.9 and 80°C in lOmin of incubation, and had the same hydrolysis pattern of larch wood xylan with the endproducts of xylobiose and xylose. Thus their activities appear essentially the same but not their stabilities.Xylanase A and B were stable from pH 2.5 to 9.0 but xylanase C was unstable above pH 5.5. Xylanase C was unstable at 70°Cwhere other two were stable.