PURIFICATION AND PARTIAL CHARACTERIZATION OF A GLYCOPROTEIN ALPHA-AMYLASE INHIBITOR FROM WHITE KIDNEY BEAN (PHASEOLUS VULGARIS. L)

Abstract

ABSTRACT A glycoprotein α-amylase inhibitor (α-AI) was extracted and purified from white kidney beans (Phaseolus vulgaris. L) by ethanol fractional precipitation, ion exchange chromatography and gel filtration column chromatography. The homogeneity of α-AI was demonstrated by gel filtration on sepharose CL-6B and SDS-polyacrylamide gel electrophoresis (SDS-PAGE). The molecular weight was estimated to be about 36,000 Da by SDS-PAGE. The glycoprotein α-AI contained 88.2% protein and was rich in aspartic acid, glutamic acid, leucine, threonine and serine, with a trace amount of cysteine. The carbohydrate moiety, determined by reversed-phase high-performance liquid chromatography (RP-HPLC), consisted of Man, Glc, Gal and Xyl in a mole ratio of 2.42:1.50:1.52:1.00. The glycan and the core protein backbone was connected by O-linkage as determined by β-elimination reaction. PRACTICAL APPLICATION The α-amylase inhibitors are important because of their potential effects of insect control and crop plant defense against pests. The selective inhibition of human α-amylase also plays an important role in reducing digestive-starch degradation in patients suffering from diabetes and has high value in controlling human weight. In this paper, an α-amylase inhibitor that effectively inhibits porcine pancreatic and salivary amylases was isolated and purified from white kidney bean. Therefore, it may have high potential pharmaceutical value as a regulative agent in reducing digestive-starch degradation in patients suffering from diabetes and obesity.