Studies on the Specificity of the Binding Site of Vicia graminea Anti‐N Lectin

Incubation of polyacrylamide gels after electrophoresis of glycoproteins with Vicia graminea lectin, labeled with radioactive iodine, showed that the lectin was bound strongly to blood‐group N and S s human sialoglycoproteins, weakly to the major sialoglycoprotein of horse red cell membranes, and was not bound to blood‐group M sialoglycoprotein. Inhibition of binding of the labeled lectin to blood‐group NN erythrocytes by untreated and modified human and horse erythrocyte glycoproteins and their structurally defined glycopeptide fragments was studied. The glycoproteins and glycopeptides were modified by desialylation, N ‐acetylation, Edman degradation or by a combination of two or more of these procedures. Desialylation and N ‐acetylation incrcased thc inhibitory activity of N and M glycoproteins and glycopeptideb. The lectin was inhibited by the NH 2 ‐terminal M and N glycopeptides of various length of the polypeptide chain, and was not inhibited by the internal tryptic glycopeptide. All modified N glycopeptides (terminated with a leucine residue) were distinctly stronger inhibitors of the lectin than respective M glycopeptides (terminated with a berine residue). In each of these two groups the most active were tryptic and pronase asialoglycopeptides with blocked amino groups. The M and N tryptic asialoglycopeptides were inactivated by Edman degradation and reactivated by the subsequent N ‐acetylation. Horse erythrocyte glycoprotein and its pronase glycopeptide were relatively weak inhibitors of V. graminea lectin, their activity was increased by desialylation, but was not affected by N ‐acetylation. Comparison of the structures of glycopeptides reacting and not reacting with V.graminea lectin suggested the following tentative conclusions. (a) V.graminea lectin can react with glycopeptides which have clusters of unsubdiluted or sialylated Gal(βl‐3)G a lNA c ‐ chains located at adjacent amino acid residues. (b) Reaction of glycopeptides with the lectin is also dependent on other (e.g. amino acid) residues present at the lectin‐binding site, namely, it is enhanced by a hydrophobic residue and is weakened by a free amino group.