title: Kinetic study of partially purified cellulase enzyme produced by Trichoderma viride FCBP-142 and its hyperactive mutants.
abstract: 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.
authors.affiliation: Institute of Mycology and Plant Pathology, University of the Punjab, Quaid-e-Azam Campus Lahore, Pakistan. drshazi81@hotmail.com
authors.forename: Shazia
authors.initials: S
authors.lastname: Shafique
authors.forename: Sobiya
authors.initials: S
authors.lastname: Shafique
mesh_terms: D002480:Cellulase / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D002482:Cellulose / Q000378:metabolism*; D004795:Enzyme Stability; D005656:Fungal Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D005818:Genetic Engineering; D006207:Half-Life; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D008970:Molecular Weight; D009154:Mutation; D011489:Protein Denaturation; D013696:Temperature; D013816:Thermodynamics; D014242:Trichoderma / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development
chemical_list: D005656:Fungal Proteins; D002482:Cellulose; D002480:Cellulase