Isolation and Characteristics of the Novel Proteinaceous Inhibitor against Pancreatic α-Amylase from White Kidney Bean ( Phaseolus vulgaris )

Abstract A novel proteinaceous α-amylase inhibitor was isolated from white kidney bean (Phaseolus vulgaris) by ammonium sulfate fractionation followed by DEAE-Sepharose Fast Flow and HiTrap DEAE-Sepharose Fast Flow chromatography. The purified inhibitor was identified as annexin (UniProt accession: V7BSM8) by HPLC-MS/MS, with a molecular mass of 35.97 kDa determined by MALDI-TOF MS. Annexin inhibited pancreatic α-amylase in a concentration-dependent manner, with an IC50 of 8.81 μM. The kinetic results were consistent with a noncompetitive inhibition pattern. Intrinsic fluorescence spectroscopy indicated that annexin binding to α-amylase induced static fluorescence quenching and conformational changes in the enzyme. Molecular dynamics simulations supported formation of a stable α-amylase–annexin complex, with electrostatic and van der Waals interactions as major energetic contributors. These findings identify annexin as a previously unrecognized α-amylase inhibitor from P. vulgaris with potential for modulating starch digestion and postprandial glycemic responses.