PJB-2019-244
Purification and characterization of cloned endo-1, 4-β-glucanase gene of Bacillus licheniformis for saccharification of plant biomass
Asma Zafar, Saima Aftab, Muhammad Nauman Aftab, Hamid Mukhtar and Mushtaq Ahmad Saleem
Abstract
Endo-1, 4-β-glucanases are the hydrolytic enzymes which are commonly used for the saccharification of plant biomass. The endo-1, 4-β-glucanase (bglC) gene of moderately thermophilic bacterium Bacillus licheniformis ATCC 14580 was successfully cloned and expressed in E. coli BL21 (DE3) strain by means of pET-22b (+) vector. Purification of recombinant endo-1, 4-β-glucanase (bglC) enzyme was done by utilizing the ammonium sulphate method followed by immobilized metal affinity chromatography (IMAC) and gel filtration. Purification fold of 10.51 was achieved along with enzyme activity and specific activity of 18.25 U/ml/min and 75 U/mg, respectively. The molecular mass of purified recombinant enzyme was 56 kDas determined by SDS-PAGE analysis. The enzyme showed resistance to temperature up to 90ºC at pH 3 - 8. The activity of enzyme was improved in the presence of metal ions particularly Co2+ and Mg2+. Organic solvents showed no significant effects on the enzyme activity and the purified enzyme also showed great stability against inhibitors such as Tween 80, β-marcaptoethanol and DTT upto 90%. Among different tested biomass, the endoglucanse enzyme showed highest saccharification (24.2%) with sugarcane bagass. These results showed that recombinant enzyme could be a useful candidate for bioconversion of natural biomass into simple sugars and eventually in biofuel production.
To Cite this article:
Zafar, A., S. Aftab, M.N. Aftab, H. Mukhtar and M.A. Saleem. 2019. Purification and characterization of cloned endo-1, 4-β-glucanase gene of Bacillus licheniformis for saccharification of plant biomass. Pak. J. Bot., 51(3): DOI: http://dx.doi.org/10.30848/PJB2019-3(27)
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