Shafique Shazia, Shafique Sobiya
Institute of Mycology and Plant Pathology, University of the Punjab, Quaid-e-Azam Campus Lahore, Pakistan.
Mikrobiologiia. 2011 May-Jun;80(3):356-65.
Cellulases are the enzymes that cleave beta-1,4 linkages of cellulose, and carbohydrate that is main part of plants' cell walls. Presently, cellulase isolation and partial purification was executed through ammonium sulfate precipitation. The isolated protein of parental and derived mutants conferred molecular weights of 30, 45 and 55 kDa. The optimum temperature for maximal cellulase activity was 50 degrees C with Ea for substrate hydrolysis of 77.73, 83.97 and 83.14 kJ mol(-1) and temperature quotient of 1.0020, 1.0022 and 1.0022 by Trichoderma viride FCBP-142, Tv-UV-5.6 and Tv-Ch-4.3, respectively. The enzyme was stable at 50 degrees C for about 60 min but rapid denaturation occurred above 55 degrees C. The enzyme showed optimum activity at pH 4.0 and involved two types of acidic and basic limbs with pKa1 and pKa2. The pKa1 of active site presented a significant shift from 2.55 to 2.9 and 3.1 by Tv-UV-5.6 and Tv-Ch-4.3, respectively in comparison to parental strain. Likewise, pKa2 moved from 6.05 to 6.5 and 6.4. Enzyme kinetics displayed Michaelis-Menten constant Km 0.6, 0.5 and 0.28 mg mL(-1) and Vmax value of 8.33, 10 and 9.09 Units mL(-1) for parental, Tv-UV-5.6 and Tv-Ch-4.3, respectively.
纤维素酶是能够切割纤维素β-1,4键的酶，而纤维素是植物细胞壁的主要成分。目前，通过硫酸铵沉淀法进行纤维素酶的分离和部分纯化。亲本菌株和衍生突变体分离出的蛋白质分子量分别为30、45和55 kDa。绿色木霉FCBP-142、Tv-UV-5.6和Tv-Ch-4.3的纤维素酶最大活性的最适温度为50℃，底物水解的活化能分别为77.73、83.97和83.14 kJ mol(-1)，温度系数分别为1.0020、1.0022和1.0022。该酶在50℃下稳定约60分钟，但在55℃以上会迅速变性。该酶在pH 4.0时表现出最佳活性，涉及两种类型的酸性和碱性基团，其pKa1和pKa2分别为2.55、2.9和3.1。与亲本菌株相比，Tv-UV-5.6和Tv-Ch-4.3活性位点的pKa1分别从2.55显著移至2.9和3.1。同样，pKa2从6.05移至6.5和6.4。酶动力学显示，亲本菌株、Tv-UV-5.6和Tv-Ch-4.3的米氏常数Km分别为0.6、0.5和0.28 mg mL(-1)，Vmax值分别为8.33、10和9.09 Units mL(-1)。
