Some factors affecting cellulase production and activity by a thermotolerant aspergillus terreus R -4 from filter mud

The growth and cellulase production by the thermotolerant Aspergillus terreus R-4 grown on filter mud [FM] waste, as the sole carbon source, were optimal when the culture medium was composed of [g/I]: FM, 25; casein, 4; KH[2]PO[4], 2; MgSO[4]. 7H[2]O, 0.5; NaCl, 2. The effect of adding various substrates in presence of FM was examined. The supplementation of glucose [5 g/I] to the fermentation medium stimulated maximal cellulase activity, but higher concentrations of glucose repressed production of cellulases. The optimal conditions for CMCase, FPase and cellobiase were at pH 4.8 and 60°C. The cellulolytic enzymes produced by A-terreus R-4 were stable over a wide range of pH values. Enzyme activity was assayed after exposure to elevated temperatures for 15 min and its thermal stability was determined

Purification and some properties of an alkaline protease from bacillus macerans

Partial purification of the crude alkaline protease produced by Bacillus macerans was carried out by fractional precipitation with acetone, ethyl alcohol or ammonium sulphate separately. The fraction obtained at 65% ammonium sulphate saturation was the most active showing 2.8-fold purification. Further purification was conducted by gel filtration on Sephadex G-100 and ion exchange chromatography of the most active protein peak on DEAE-cellulose. A pure alkaline protease enzyme was obtained [molecular weight 24000]. Maximum activity of the pure enzyme was obtained at a protein concentration of 25 micro g/reaction mixture, and optimum substrate concentration was 10 mg casein/reaction mixture. The amino acid composition of the pure enzyme was similar to that of other bacterial alkaline protease. The optimum temperature of the reaction was 50°C and the optimum pH value was 10.8. The enzyme was fairly stable to heat treatment and retained 56.5% of its activity after heating for 15 min at 60°. Ca [2+] ions slightly activated the enzyme, while Co[2+], Ba[2+], Fe[3+]and EDTA partially inhibited the enzyme activity. Cu[2+], Hg[2+]and Zn[2+]strongly inhibited the enzyme activity. Phenylmethylsulforyl fluoride [PMSF] inhibited the enzyme suggesting the presence of serine in the active site of the enzyme

Purification and characterization of chitinase produced by bacillus amyloliquefaciens

The crude chitinase preparation obtained from Bacillus amloliquefaciens cultures was partially purified by fractional precipitation with ethanol, acetone or ammonium sulphate. The 65% ammonium sulphate fraction was the most active and was further purified by gel filtration on Sephadex G-200 followed by ion exchange chromatography on Ecteola ET-11 cellulose yielding 4 chitinolytic enzyme components. Two enzyme components, CHII and CHIII, showed high activity and protein recovery. The purity of both enzymes was checked by polyacrylamide gel electrophoresis. CHII enzyme was more specifid for chitin as substrate and showed a higher thermostability than CHIII. Both enzymes showed a temperature optimum of 45°C and pH optimum of 5.5 to 5.9. They did not show specific cationic requirements and were partially inhibited by zinc, cupric, silver and cobalt ions. Their amino acid compositions were different. SDS electrophoresis revealed that each enzyme was formed of 2 subunits of different molecular weights, CHII subunit had a M.W. of 26.5 and 23.5 KD while CHIII was 31.6 and 27.5 KD