Purification, biochemical characterization and Insilico modeling of α-amylase from Vicia faba

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.