Isolatton Of High Efficiency Cellulose-Degrading Penicillium Citrinum And Its Enzymology Characteristics

A new strain of high efficiency cellulose-degrading is isolated from soil of Yun Nan province. Morphological and physiological properties of this strain have been studied.According to these results,this strain was identified as Penicillium citrinum,named CR-2..The medium components of the optimal liquid state fermentation of CR-2,the purification of endoglucanase from CR-2 and characteristics of the purified endoglucanase are studied.The results are illustrated as follows:1.CR-2 strain is cultivated by cellulose plate inversely at constant temperature28℃,after 3days cultivation,the green and gray colonies appeared.After dying by Congo Red solution and decoloring by NaCl solution,the transparent hydrolysis circle appeared.To see the characterization of the colonies,the mycelia and conidia,strain is inoculated on the PDA medium after isolation.The colonies are white at first then turn to green and gray like floccus blanket(Fig2-2),grow up slowly. Cultivate CR-2 strain at 30℃10days,the diameter of colonies are less than 2cm and the back of the colonies are yellow.Mycelia are no colour and the width is 0.8～1.4μm.Peduncles of the conidia are short,like pillars and have ramifications,the length is 5～15μm and the width is 1.0～1.5μm;The below parts of peduncles have no ramifications,and have more than 3～5 conidia,3.1 7.1×1.6～2.8μm like flasks,the widest are near the basic,the width of top is 0.5～1.2μm;Conidia are like rotundity and dispersive.According to ribosome-DNA ITS region sequence comparing,the strain is identified as Penicillium citrinum.2.CR-2 strain is inoculated in different carbon source and nitrogen source culture mediums.CMCase in each medium and are compared so that the optimal liquid state fermentation medium is decided.The results show that optimum conditions of producing cellulase are as follows: The temperature is 30℃,the initial pH,the shaking speed is 200r/min,inoculation amount is 10%, cellulase production,highest cellulase activities are obtained by using maize staff carbon sources. Under the conditions mentioned above,the activities of CMCase and Fpase,β-glucosidase in cultivation broth are 49.91IU/ml and 26.9IU/ml,0.12IU/ml,respectively. 3.The crude cellulase broth of CR-2 strain is purified by ammonium sulfate fractionation,dialysis and Sephadex G-75,DEAE-PE52 column chromatography to obtain a purified component.Meanwhile,this shows molecular weights of this enzyme about 27.26kDa..4.The optima degrading conditions for cellulose obtained are pH4 and 70℃.The enzyme is stable between pH3 and 6,and at the temperature below 70℃,The enzyme activity is distinctive inhibited by metal ions such as Cu²⁺,Mn²⁺,Ag⁺ but enhanced by Na⁺,K⁺.The Vmax and Km of endoglucanase are Vmax=2.88 mg/min.ml and Km=0.046 mg/ml.