Cloning and overexpression of raw starch digesting α-amylase gene from Bacillus subtilis strain AS01a in Escherichia coli and application of the purified recombinant α-amylase (AmyBS-I) in raw starch digestion and baking industry
| Autor: | Ashis K. Mukherjee, Anjan Borah, Jetendra Kumar Roy, Charu Lata Mahanta |
|---|---|
| Rok vydání: | 2013 |
| Předmět: |
biology
Molecular mass Starch Process Chemistry and Technology food and beverages Bioengineering Maltose Bacillus subtilis medicine.disease_cause biology.organism_classification Biochemistry Catalysis Hydrolysis chemistry.chemical_compound chemistry medicine biology.protein Extracellular Amylase Escherichia coli |
| Zdroj: | Journal of Molecular Catalysis B: Enzymatic. 97:118-129 |
| ISSN: | 1381-1177 |
| Popis: | Considering the economic and industrial relevance of α-amylases used in food and starch industries, a raw starch digesting α-amylase gene ( amyBS-I ) from Bacillus subtilis strain AS01a was cloned and expressed in Escherichia coli BL21 cells. The gene also includes its signal peptide sequence (SPS) for facilitating the efficient extracellular expression of recombinant α-amylase (AmyBS-I) in correctly folded (enzymatically active) form. The native AmyBS-I consists of 659 amino acids with a molecular mass and p I of 72,387 Da and 5.8, respectively. The extracellular secretion of AmyBS-I after response surface optimization of culture conditions was found to be 7-fold higher as compared to its production under non-optimized conditions. Purified AmyBS-I demonstrated optimum activity at 70 °C and pH 6.0. It shows K m and V max values toward soluble starch as 2.7 mg/ml and 454 U/ml, respectively. Further, it does not require Ca 2+ ion for its α-amylase activity/thermo-stability, which is an added advantage for its use in the starch industry. The AmyBS-I also hydrolyzed a wide variety of raw starches and produced maltose and glucose as main hydrolyzed products. The bread dough supplemented with AmyBS-I showed better amelioration of the bread quality as compared to the bread supplemented with commercial α-amylase. |
| Databáze: | OpenAIRE |
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