Cellulase-producing Fermentation Conditions Of Penicillium Purpurogenum HBZ003 From Mangrove And Properties Of Enzymes

The cellulase-producing fermentation conditions, including cellulase-producing media and its culturing conditions, of Penicillium purpurogenum HBZ003 from mangrove soil were firstly studied, and then the enzyme of the strain was extracted and purificated, subsequently the properties of the enzymes were investigated.The optimum fermentation conditions of Penicillium purpurogenum HBZ003 were as follows: wheat bran 16 g/L, CMC 4 g/L, (NH,)2SO45 g/L, KH2PO43 g/L, NaCl 6g/L, CaCl20.5 g/L, Tween-20 0.5Ml/L, initial pH 4.0,10%(v/v) of the seed was inoculated in 250mL flask containing 100mL broth, and incubated for 5days at 30℃,160r/min,and CMCase and FPA reached 16.04U and 4.08U.To minimize the mixed protein, the range of ammonium sulfate saturation 20 to 60% was selected, so that the cellulase could be fully purified; By the purification steps of ammonium sulfate fractionation, dialysis and Sephadex G-100 chromatography,and so on, two kind of components of Penicillium purpurogenum HBZ003 cellulase system with endo-glucanase activity and P-glucosidase activity were isolated and purified, respectively, which purified enzyme specific activity were increased to 6.52 and 19.30 and the recoveries were 21.79%and 28%;Two tubes of enzyme liquid with Cx and BG activity after purification were run with SDS-PAGE, showing multi-band, which demonstrated that the liquid were composed of three Subunits or three kinds of proteins of Cx and BG enzyme respectively.The molecular weight of the three were 80kD,65kD,55kD and 150kD,120kD,100kD,respectively; The Km value of Cx enzyme was 2.03×10-2g/mL, Vmax was 0.20mg/(min.mL), BG's Km value was 4.23×10-3g/mL, Vmax was 0.37mg/(min.mL); The activity of Cx enzyme is high between 35℃-70℃,while BG enzyme is between 55℃-70℃with high activity; The Cx and BG activity could maintain more than 90% after insulating 0.5h in the temperature range of 30-55℃; The optimal pH ranges of Cx and BG were 3 to 5 and 3.5-6,respctively; Mn2+, Ca2+, Cu2+, Fe2+, Zn2+ activated the Cx enzyme, While Mg2+, Co2+ inhibited it. Mn2+, Cu2+, Fe2+, Zn2+ activated the BG enzyme, while Ca2+, Mg2+, Co2+ inhibited it.