Purification, biochemical characterization and Insilico modeling of α-amylase from Vicia faba
| Autor: | Vinay Kumar Singh, Fatima Ahmad, Arvind M. Kayastha, Kritika Singh, Kanwal Kayastha |
|---|---|
| Rok vydání: | 2017 |
| Předmět: |
0301 basic medicine
chemistry.chemical_classification 030102 biochemistry & molecular biology biology Molecular mass Chemistry food and beverages Condensed Matter Physics Atomic and Molecular Physics and Optics Electronic Optical and Magnetic Materials Conserved sequence Vicia faba 03 medical and health sciences 030104 developmental biology Enzyme Biochemistry Protein purification Materials Chemistry biology.protein Amylase Homology modeling Physical and Theoretical Chemistry Binding site Spectroscopy |
| Zdroj: | Journal of Molecular Liquids. 234:133-141 |
| ISSN: | 0167-7322 |
| Popis: | α-Amylase from germinated broad bean (Vicia faba) was purified 2050 fold to electrophoretic homogeneity with a final specific activity of 369 U/mg. On SDS-PAGE the purified enzyme showed a single band with a molecular mass of 45 kDa. MS/MS analysis of the above sample confirmed the presence of α-amylase in the purified sample. Optimal activity of this enzyme was observed at pH 6.0 and 65 °C with activation energy of 8.47 kcal/mol. Km for hydrolysis of starch was found to be 4.6 mg/mL. Further characterization using Insilico studies (InterProScan and phylogenetic approach) on broad bean α-amylase identified it to contain essential domains and conserved sequences belonging to α-amylase family. Homology based significant quality 3D model was generated showed similar binding site residues with other conserved α-amylase domain using CDD blast. Substrate acarbose complexed with broad bean α-amylase complied with the residues participating in the interaction thereby indicating its proper folded structure. |
| Databáze: | OpenAIRE |
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