Sperm-egg binding in the sea urchin: a high level of intracellular ATP stabilizes sperm attachment to the egg receptor.

Previous studies have established that a recombinant protein fragment (45A) of the egg receptor for sperm of the sea urchin Strongylocentrotus purpuratus exhibits several characteristics that are consistent with that expected of a receptor. Using a quantitative sperm binding assay with glutathione S-transferase fused to a recombinant protein containing the C-terminal half of the 45A construct immobilized on glutathione beads, it was found that the interaction between sperm and this protein is a kinetically transient event. Sperm binding to the receptor fragment reached a maximum at 20 s after adding sperm in the presence of egg jelly to beads coated with recombinant receptor. In the next 20-120 s, approximately 50-70% of the sperm detached from the beads. Similar phenomena were observed when the kinetics of sperm binding to dejellied, glutaraldehyde-fixed eggs were studied. Because the acrosome reaction, a prelude to binding, is known to be accompanied by a decrease in the ATP level of sperm, we studied the effect of various inhibitors on both sperm detachment and the level of ATP. It was found that the detachment rate increased slightly when respiration inhibitors that blocked ATP production in mitochondria were added. In contrast, the dynein ATPase inhibitor, erythro-9-[3-hydroxynonyl]adenine, which is known to inhibit flagellum motility by blocking ATP utilization, stabilized the binding of sperm to the receptor and allowed maintenance of a high internal ATP level. Immotile, tailless sperm that physically lacked dynein ATPase, and therefore sustained their internal ATP levels, also exhibited stable binding provided that the sperm and beads were physically mixed. These results suggest that the internal ATP level of the sperm controls the stability of its binding to the receptor. The possible mechanism of the detachment and its significance with respect to the overall process of fertilization are discussed.