Sea urchin egg receptor for sperm: the oligosaccharide chains stabilize sperm binding

Sulfated O-linked oligosaccharides from the sea urchin egg receptorhave been shown to bind to acrosome-reacted sperm and to inhibitfertilization in a competitive bioassay. However, the inhibitoryactivity of these isolated chains was much lower than that ofa recombinant protein representing a portion of the extracellulardomain of the receptor. Because the isolated oligosaccharideslacked the potential polyvalency that they might have when linkedto the polypeptide backbone, in the current study we asked iftheir inhibitory activity could be increased by chemically couplingthem to a protein to form a neoglycoprotein. Using a recombinantfragment of the receptor we could not detect an oligosaccharidedependent increase in inhibitory activity with this neoglycoprotein,probably because of the much higher inhibitory activity of thepolypeptide backbone. Therefore, we examined the activity ofthe oligosaccharides coupled to a protein lacking the abilityto inhibit fertilization, namely, bovine serum albumin. A markedincrease in the inhibitory activity of the oligosaccharideswas observed with this neoglycoprotein. Finally, because inhibitionby the oligosaccharides and the polypeptide was measured inan end point assay, namely, inhibition of fertilization, wesought a more direct, kinetically sensitive way to measure theirproperties. Accordingly, an assay was devised (R.L.Stears andW.J.Lennarz, unpublished observations) involving measurementof sperm binding to beads that was dependent on the presenceof the receptor or its components. This assay revealed thatsperm binding to beads via the recombinant protein peaked at10 sec and then declined. In contrast, binding mediated by neoglycosylatedrecombinant protein reached a plateau. Thus, binding of spermto the oligosaccharides resulted in a more stable interactionthan that observed in binding to the polypeptide backbone. sperm binding sea urchin egg sperm binding oligosaccharide