Isolation of the lectin and an L4 isolectin from Phaseolus vulgaris by affinity chromatography on insoluble ovomucoid
Gonçalves, R.B.; Costa, C.P.
Brazilian Journal of Medical and Biological Research 28(2): 191-194
1995
ISSN/ISBN: 0100-879X PMID: 7581040 Document Number: 445534
Lectins from extracts of Phaseolus vulgaris seeds have potent cell-agglutinating and lymphocyte-stimulating activity. An affinity adsorbent for lectins with specificity for the oligosaccharide structure was prepared by transforming ovomucoid, an oligosaccharide-rich glycoprotein, into an insoluble and stable gel. The ovomucoid was made insoluble by boiling a 20% solution (200 mg/ml) in 0.1 M Tris-HCl, pH 8.9, for 20 min. This insoluble gel was desialylated by treatment with 50 mN sulfuric acid for 1 h at 90 degree C and fixed with 1% glutaraldehyde, pH 7.4, for 10 min. The Phaseolus lectin and the L-4 isolectin could be isolated essentially in a single-step procedure, using different eluting conditions: 50 mM sodium formate buffer, pH 3.0, was used for PHA elution; a different column was eluted with 15 mM sodium tetraborate, pH 8.0, for desorbed L-4 isolectin. Polyacrylamide gel electrophoresis of the lectin showed five distinct bands, whereas the L-4 isolectin only presented one band. From 250 mg of saturated column, 8.25 mg of PHA was isolated. This adsorbent could be used several times with little change in binding capacity or selectivity.