PURIFICATION AND PROPERTIES OF A MITOGENIC LECTIN FROM WISTARIA-FLORIBUNDA SEEDS

Extracts of Wistaria floribunda seeds contain separable erythroagglutinating and lymphocyte mitogenic activities. Cheung et al. [Cheung, G., Haratz, A., Katar, M., Skrokov, R., & Poretz, R. D. (1979) Biochemistry 18, 1646] have reported a method of purifying the agglutinin. The mitogen as reported here is purified by a compatible procedure. The protein of an aqueous extract of ground seeds precipitated by ethanol contains erythroagglutinating and mitogenic activities. Following dissolution of the precipitate, the erythro-agglutinin is adsorbed onto polyleucyl hog gastric mucin (allowing its purification by desorption with a saccharide inhibitor, lactose), resulting in a solution with a trace of erythroagglutinating activity and much of the mitogenic activity. After isoelectric precipitation of the residual erythroagglutinin and other contaminants, the mitogenic activity is further purified by ion-exchange chromatography. The mitogen is obtained in a homogeneous state by liquid isoelectric focusing. The purified mitogen is isoelectric at pH 7.3. It displays a molecular weight of 66 000 by gel filtration and possesses two sodium dodecyl sulfate dissociable subunits of 32000. The purified mitogen exhibits one protein and carbohydrate staining band on both basic and acidic polyacrylamide gel electrophoresis. The mitogen is approximately 10% neutral carbohydrate by weight and has no detectable erythroagglutinating activity. It is mitogenic for murine and human lymphocytes. Binding of the mitogen to the lymphocyte cell surface is inhibited by d-mannose and thyroglobulin glycopeptides. © 1979, American Chemical Society. All rights reserved.