Purification and biochemical properties of a glucose-stimulated β-D-glucosidase produced by <Emphasis Type="Italic">Humicola grisea</Emphasis> var. <Emphasis Type="Italic">thermoidea</Emphasis> grown on sugarcane bagasse

The effect of several carbon sources on the production of mycelial-bound β-glucosidase by Humicola grisea var. thermoidea in submerged fermentation was investigated. Maximum production occurred when cellulose was present in the culture medium, but higher specific activities were achieved with cellobiose or sugarcane bagasse. Xylose or glucose (1%) in the reaction medium stimulated β-glucosidase activity by about 2-fold in crude extracts from mycelia grown in sugarcane bagasse. The enzyme was purified by ammonium sulfate precipitation, followed by Sephadex G-200 and DEAE-cellulose chromatography, showing a single band in PAGE and SDS-PAGE. The β-glucosidase had a carbohydrate content of 43% and showed apparent molecular masses of 57 and 60 kDa, as estimated by SDS-PAGE and gel filtration, respectively. The optimal pH and temperature were 6.0 and 50°C, respectively. The purified enzyme was thermostable up to 60 min in water at 55°C and showed half-lives of 7 and 14 min when incubated in the absence or presence of 50 mM glucose, respectively, at 60°C. The enzyme hydrolyzed p-nitrophenyl-β-D-glucopyranoside, p-nitrophenyl-β-Dgalactopyranoside, p-nitrophenyl-β-D-fucopyranoside, p-nitrophenyl-β-D-xylopyranoside, o-nitrophenyl-β-Dgalactopyranoside, lactose, and cellobiose. The best synthetic and natural substrates were p-nitrophenyl-β-Dfucopyranoside and cellobiose, respectively. Purified enzyme activity was stimulated up to 2-fold by glucose or xylose at concentrations from 25 to 200 mM. The addition of purified or crude β-glucosidase to a reaction medium containing Trichoderma reesei cellulases increased the saccharification of sugarcane bagasse by about 50%. These findings suggest that H. grisea var. thermoidea β-glucosidase has a potential for biotechnological applications in the bioconversion of lignocellulosic materials.     

Composition of cholesterol esters in blood plasma of mouse, rat and rabbit

Mixtures of cholesterol estes were extracted from the blood plasma of mouse (C3H/He, C57BL/6J and Jcl : ICR), rat (Wistar) and rabbit (JW-NIBS/Rabiton) and purified by thin-layer chromatography. Fatty acid moieties of the esters were methylated with BF3-methanol and analyzed quantitatively by gas chromatography. Although linoleate was most abundant (over 30% of the total esters) and myristate was found only in traces, the composition of the esters varied markedly among the species. Mice had 15.3 to 17.4% of arachidonate, but only a trace of linolenate and no stearate. In rats, arachidonate and linoleate were found in almost equal quantities, but stearate, linolenate and docosahexaenoate were present in only trace amounts. Rabbits had more palmitate and oleate and less arachidonate and eicosapentaenoate than the other two animals, but no docosahexaenoate.     

A Novel Molybdenum Protein with a Low Molecular Mass in Rhodobacter sphaeroides f. sp. denitrificans

A novel molybdenum cofactor-containing protein with a low molecularmass of 20 kDa was found in a photodenitrifier, Rhodobactersphaeroides f. sp. denitrificans. The protein was located inthe cytoplasm and was produced constitutively. (Received March 25, 1993; Accepted May 14, 1993)