Purification and characterization of a lectin from Wistaria floribunda seeds.
A lectin from Wistaria floribunda seeds which specifically binds to N-acetyl-D-galactosamine was purified to homogeneity as judged by polyacrylamide gel electrophoresis. Its molecular weight was estimated to be 68,000 by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. It dissociated into subunits on reduction with 2-mercaptoethanol with concomitant loss of hemagglutinating activity. On oxidation in air, the subunits reassociated into the lectin molecules with hemagglutinating activity. Carboxymethylation of the subunits with iodoacetic acid prevented their reassociation on oxidation in air. The molecular weight of the subunits was 32,000, which is about one-half that of the native lectin, suggesting that the lectin consists of two subunits. The results total, NH2-terminal, and COOH-terminal amino acid analyses, and mapping of the tryptic digest of the lectin indicated that these two subunits were indistinguishable and were probably identical, and that they were linked together covalently through a single disulfide bond. Equilibrium dialysis experiments show that the lectin and its subunit molecules are divalent and monovalent, respectively, with respect to sugar binding. The lectin is a glycoprotein, containing 3.2% carbohydrate. The carbohydrate moiety is composed of mannose, galactose, and glucosamine is a molar ratio of 1:2:1 and these sugars seem to be linked as a single oligosaccharide chain to each subunit of the protein.
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D005944:Glucosamine / Q000032:analysis; D006386:Hemagglutination Tests; D006601:Hexoses / Q000032:analysis; D006801:Humans; D037102:Lectins / Q000302:isolation & purification; D046911:Macromolecular Substances; D008970:Molecular Weight; D010084:Oxidation-Reduction; D010446:Peptide Fragments / Q000032:analysis; D037121:Plant Lectins; D012639:Seeds / Q000032:analysis*; D013045:Species Specificity; D014357:Trypsin
D000596:Amino Acids; D006601:Hexoses; D037102:Lectins; D046911:Macromolecular Substances; D010446:Peptide Fragments; D037121:Plant Lectins; D014357:Trypsin; D005944:Glucosamine