Studies On Screening For Cellulose-degrading Actinomyces And Its Functional Identification

In this study, a cellulose-degrading Actinomyces called SD1-4 screened from different habitats was identified as Streptomyces (Streptomyces sp.) based on the result of morphological, physiological and molecular studies.Liquid fermentation conditions of the strain SD1-4 were optimized for enzyme production, the results show that the initial pH was 8.0,, and optimum temperature was set at 39℃, and the optimum nitrogen source was yeast extract, the best salt addition level was 60 mgKCI and 15mg CaCl2, the best Sterilization time 40 min, the optimum liquid volume 50ml/250ml. The result also indicated that 0.15% Triton X-100 inhibit the enzyme production, reverse was true at 0.05% Triton X-100 treatment , PEG8000 and Tween -20 at two treatment concentrations showed promotion on the enzyme, however, SDS showed inhibitory effect on it; for enzyme production reached to the peak.detected at the first 9 daysThe result also showed Pennisetum purpereum Schumach was the most suitable carbon source; the index of selection energy grass was high in the contents of fiber, sugar , and proteins but low in the contents of amylose, glucose and fructose. were relatation analysis, principal component analysis, regression analysis and cluster analysis were used as the optimal statistical tools, but gray correlation analysis and path analysis were not suitable for screening. The nutrient content (9.04% soluble sugar, soluble protein was 3.37%, the starch content of 5.95%, the crude fiber content of 65.95%) of Pennisetum purpereum Schumach was the best fermentation ingredients.Designed by Box-Bohnken a related model has been constructed between cellulase production (endogulucanase, exoglucanase and glucosidase) and seven factors (temperature, pH, particle size, peptone concentration and humidity, inoculum and time). The difference of three models (GA-ANN, ANN and RSM) in optimization was compared. The R values were 0.869, 0.906 and 0.867 in the optimum conditions, compared to the R values of 0.796, 0.881 and 0.840 in non-optimal conditions ,, indicating that GA-ANN showed thw best in capturing nonlinear behavior.A kind of endoglucanase from the extracellular crude enzyme of SD1-4 was purified and concentrated by PEG, ammonium sulfate precipitation, ethanol washing, Sephadex G-200, DEAE A-50 and Sephadex G-200. The enzyme was monomer with a molecular weight, about 38KD and visuated in a band in SDS-PAGE. It could hydrolyze CMC-Na, and the optimum temperature was 60℃and pH 7.0. Vmax and Km, were 22.57 U·ml-1·min-1 and 0.14%. respectively. It wass found that 0.2 mM Co2+ added to media had a strong role in promoting enzyme,activity, but 0.8 mM Co2+ showed inhibitory effect on the enzyme activity.