Purification and characterisation of a cellulase obtained from cocoa (Theobroma cacao) pod-degrading Bacillus coagulans Co4

Abstract

Objective: Some properties of cellulase purified from the culture supernatant of Bacillus coagulans Co4, isolated from cocoa pod dumpsite were investigated for possible biotechnological applications. Methods: The crude cellulase was purified to apparent homogeneity using a combination of acetone precipitation, CM Sepharose CL-6B ion exchange chromatography and gel filtration on Sephadex G-100. The molecular and thermodynamic properties of the purified enzyme were studied following standard procedures. Results: The specific activity of the purified cellulase rose from 0.10 to 47 units/mg of protein, at the end of purification. The molecular weight was found to be 14.5 kDa; and an apparent K m value of 0.18±0.06 mg/ml of carboxylmethylcellulose. The optimum pH and temperature were 7.5 and 60 o C respectively. The activation energy for carboxylmethylcellulose hydrolysis (E a ) was 16.5 kJ/mol. Na+ and K+ had no effects on its activity at concentrations up to 200 mM, whereas Ca 2+ and Mg 2+ served as inhibitors at concentrations above 25 and 40 mM respectively. The cellulase retained 40% residual activity when heated at 60oC for 40 minutes. Conclusison: On the basis of these properties, it is concluded that the purified cellulase is moderately thermostable and may have applications in the bioconversion of agricultural wastes into economically useful products.