Purification and characterization of an α-galactosidase from Phaseolus coccineus seeds showing degrading capability on raffinose family oligosaccharides.

An acidic α-galactosidase (EC 3.2.1.22) designated as Phaseolus coccineus seeds galactosidase (PCG) was purified from P. coccineus seeds using ion-exchange chromatography on DEAE- and CM-cellulose, Q- and SP-Sepharose and gel filtration on Superdex 75. The molecular weight of PCG was 43 kDa as judged by SDS-PAGE and gel filtration. Two inner peptides of PCG were sequenced by MALDI-TOF-MS. The optimum pH and temperature was 3.0 and 70 °C, respectively but was stable up to 60 °C for 30 min. The enzyme activity was inhibited by NBS signifying the pivotal role played by tryptophan in the catalytic activity of the enzyme. The Km for hydrolysis of pNPGal was 0.0025 mM. Besides hydrolyzing pNPGal, α-galactosidases also hydrolyzed natural substrates such as melibiose, raffinose and stachyose. Hence it can be exploited commercially for improving the nutritional value of soymilk. Thus the PCG has great potential in the feed industries for removal of non-digestible oligosaccharide from legumes.