Carbohydrate Specificity of Sea a Cell Surface Lectin Mediating Urchin Sperm Bindin: Sperm-Egg Adhesion

We have examined the carbohydrate specificity of bindin, a sperm protein responsible for the adhesion of sea urchin sperm to eggs, by investigating the interaction of a number of polysaccharides and glycoconjugates with isolated bindin. Several of these polysaccharides inhibit the agglutination of eggs by bindin particles. An egg surface polysaccharide was found to be the most potent inhibitor of bindin-mediated egg agglutination. Fucoidin, a sulfated fucose heteropolysaccharide, was the next most potent inhibitor, followed by the egg jelly fucan, a sulfated fucose homopolysaccharide, and xylan, a fl(1--,4) linked xylose polysaccharide. A wide variety of other polysaccharides and glycoconjugates were found to have no effect on egg agglutination. We also report that isolated bindin has a soluble lectinlike activity which is assayed by agglutination of erythrocytes. The bindin lectin activity is inhibited by the same polysaccharides that inhibit egg agglutination by particulate bindin. This suggests that the egg adhesion activity of bindin is directly related to its lectin activity. We have established that fucoidin binds specifically to bindin particles with a high apparent affinity (Kd = 5.5 X 10 -8 M). The other polysaccharides that inhibit egg agglutination also inhibit the binding of 1281-fucoidin to bindin particles, suggesting that they compete for the same site on bindin. The observation that polysaccharides of different composition and linkage type interact with bindin suggests that the critical structural features required for binding may reside at a higher level of organization. Together, these findings strengthen the hypothesis that sperm-egg adhesion in sea urchins is mediated by a lectin-polysaccharide type of interaction. A growing body of evidence indicates that carbohydrates of the cell surface serve as specific recognition determinants in intercellular adhesion phenomena (5, 13). Sea urchin fertilization represents an example of specific ceil recognition and adhesion, the outcome of which is the fusion of the gametes and subsequent activation of the zygote. Sperm adhesion to the egg vitelline layer is mediated by the protein bindin, the major component of the sperm acrosome granule, which is exposed by exocytosis during the acrosome reaction (29, 32, 33). Two lines of evidence support the hypothesis that bindin mediates sperm-egg adhesion. Immunocytochemistry with anti-bindin localizes bindin to the site of sperm-egg adhesion (19), and insoluble bindin particles agglutinate sea urchin eggs species specifically (6, 8). This specificity of egg agglutination by bindin corresponds to the specificity observed for sperm adhesion. The available evidence suggests that egg surface glycoconjugates serve as receptors for bindin (7-9, 32). The egg surface glycoconjugate implicated in the adhesion of sperm to eggs is a high molecular weight polysaccharide containing fucose, xylose, galactose, and glucose (9). In a preliminary communication, Vacquier and Moy (34) reported that bindin agglutinates erythrocytes and that the hemagglutination activity is inhibited by several simple sugars. We have not been able to consistently reproduce this observation with particulate bindin; however, we have found that bindin suspensions contain a soluble hemagglutination activity that is reproducibly inhibited by simple sugars. Thus bindin satisfies the classical definition of a lectin; it agglutinates erythrocytes and this agglutination is inhibited by specific sugars (12). We have further defined the carbohydrate-binding specificity of bindin in this report by using a series of polysaccharides and neoglycoproteins. The results of these investigations demonstrate that the egg agglutinating activity of bindin is directly related to its lectin activity. THE JOURN,',L OF CELL BIOLOGY • VOLUME 94 IUtV 1982 123-128 © The Rockefeller University Press • 0021-9525/82/07/0123/06 gl.00 1 23 on A ril 4, 2017 D ow nladed fom Published July 1, 1982