Production,purification and characterization of a novel GH 12 family endoglucanase from Aspergillus terreus and its application in enzymatic degradation of delignified rice straw |
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Institution: | 1. Departamento de Física e Informática, Instituto de Física de São Carlos, Universidade de São Paulo, São Carlos, SP, Brazil;2. Departamento de Biotecnologia, Escola de Engenharia de Lorena – Universidade de São Paulo, Lorena, SP, Brazil;3. Laboratório Nacional de Ciência e Tecnologia do Bioetanol, Centro Nacional de Pesquisa em Energia e Materiais, Campinas, SP, Brazil;4. Laboratório Nacional de Biociências, Centro Nacional de Pesquisa em Energia e Materiais, Campinas, SP, Brazil |
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Abstract: | Endoglucanase production was carried out using in-house isolate Aspergillus terreus on rice straw under solid state fermentation. An increase of 1.25-fold endoglucanase production was obtained under optimized conditions using response surface methodology. The enzyme was purified to homogeneity by gel filtration chromatography. Its molecular weight was determined as 28.18 kDa by gel filtration and 29.13 kDa on SDS-PAGE. The enzyme displayed maximum activity at 50 °C and pH 4.8. It was stable for 240 min at 50 °C and 120 min at 60 °C but rapidly inactivated at 70 °C. The purified enzyme was specific towards carboxymethyl-cellulose but showed no activity for cellobiose or xylan. Maximum velocity (Vmax) and KM were 16.15 μmol min−1 mg−1 and 12.01 mg ml−1, respectively. AgNO3, KCl, NaCl, and MnSO4 were found to inhibit enzyme activity while CaCl2 and ZnSO4 activated the enzyme. Internal peptide mass fingerprinting analysis identified that the protein belongs to GH12 superfamily endoglucanases. External supplementation of the purified enzyme to the crude cellulase showed 38.7% increase in saccharification efficiency of the delignified rice straw compared to the crude cellulase alone. The results demonstrated that the addition of GH 12 family purified endoglucanase to the crude cellulase can efficiently convert lignocellulosic biomass to fermentable sugars. |
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Keywords: | Endoglucanase Purification Gel filtration Kinetics Enzymatic hydrolysis |
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