Purification and Characterization of Four Isolectins of Mushroom(Agaricus bisporus)

Abstract

Four lectins were purified from a mushroom (A. bisporus) by ammonium sulfate fractionation, anion-exchange chromatography, affinity chromatography on bovine submaxillary mucin-Sepharose 4B and preparative isoelectric focusing. They were designated as ABA-I (pI 6.70), II (pI 5.98), III (pI 5.69) and IV (pI 5.53). Polyacrylamide gel electrophoresis of each lectin in the presence of sodium dodecyl sulfate gave a single band with an apparent MW of 16,000 daltons. Sedimentation equilibrium analysis suggested that each lectin is a tetramer of subunits. The 4 lectins have quite similar carbohydrate-binding specificities. The hemagglutination activities of the lectins were effectively inhibited by bovine and porcine submaxillary mucins (BSM and PSM), and NH2-terminal glyco-octapeptides obtained by cyanogen bromide cleavage of human erythrocyte glycophorin A. In addition, desialylated PSM-glycopeptides were more potent inhibitors than untreated PSM-glycopeptides. Among monosaccharides and their glycosides, only methyl N-acetyl-.alpha.-galactosaminide inhibited lectin binding at a high concentration, but a synthetic oligosaccharide, O-.beta.-galactopyranosyl-(1 .fwdarw. 3)-O-(2-acetamido-2-deoxy-.alpha.-D-galactopyranosyl)-N-tosyl-L-serine, was a strong inhibitor.