THE INVOLVEMENT OF O-LINKED OLIGOSACCHARIDE CHAINS OF THE SEA-URCHIN EGG RECEPTOR FOR SPERM IN FERTILIZATION

Recent investigations on the sea urchin egg receptor for sperm have led to its sequencing and the demonstration that it is a 350 kDa glycoprotein, In the current study, the N- and O-linked oligosaccharide chains were cleaved from the protein fractionated on concanavalin A-agarose, The putative O-linked oligosaccharide chains that did not bind to the lectin were further fractionated by anion-exchange chromatography. Using a competition bioassay that measured the ability of these oligosaccharide chains to inhibit fertilization, it was found that the N-linked chains were devoid of inhibitory activity, Rather, the inhibitory activity was localized to the O-linked chains, with the most highly charged, sulphated chains showing the highest inhibitory activity, The bioactive oligosaccharides were labelled by reduction and assayed for binding to sperm. The results of the binding assay, coupled with the fertilization bioassay, indicate that the oligosaccharides inhibit fertilization by binding to acrosome-reacted sperm, The bioactive oligosaccharide lacked species specificity in fertilization bioassays, unlike the intact receptor and a recombinant aglyco protein containing only the extracellular domain of the receptor. Since previous work showed that the recombinant protein inhibits fertilization species specifically and binds to acrosome-reacted sperm, a two-step model of sperm-egg interaction is proposed, The first step is postulated to be a low-affinity ionic interaction of the sulphated O-linked oligosaccharide chains of the receptor with sperm that is not species specific, This is followed by a high-affinity, species-specific interaction of the sperm with one or more binding sites on the polypeptide chain of the receptor.