Binding of concanavalin A to the surface of sea urchin eggs and its alteration upon fertilization.

The binding of concanavalin A (Con A) to Strongylocentrotus purpuratus eggs has been investigated using 125I-concanavalin A (125I-Con A). The lectin binds specifically to the egg surface and does not produce agglutination of the eggs. High 125I-Con A concentrations are necessary to saturate all of the surface binding sites. Scatchard plots of the binding data are biphasic and may be interpreted as showing two main classes of sites. Unfertilized eggs have 4 X 10(8) high affinity sites/egg (Ka=8 X 10(-7) M) and 4.5 X 10(8) istes of lower affinity (Kb=4 X 10(-6) M). The sites may be assigned to different layers of the egg surface by studying the effects of removal of egg coats on the binding reaction. Removal of the jelly coat by washing eggs at pH 4.5 has no effect on binding. However, disruption of the vitelline layer with dithiothreitol leads to a decrease in the number of high affinity sites. After fertilization, the high affinity sites are found upon the fertilization membrane produced from the vitelline layer. Low affinity sites predominate in the plasma membrane, and are not affected by treatments which alter the vitellinelayer. The number of low affinity sites double upon fertilization, with the insertion of new membrane into the egg surface, as a result of cortical granule exocytosis. The doubling of sites is not due to hyaline material released from the cortical vesicles at fertilization, and thus these sites appear to reside upon the new membrane that is inserted from the cortical vesicles. If eggs are activated with ammonia, bypassing the cortical reaction, no change in the binding of ConA occurs. Con A inhibits fertilization at concentrations higher than 0.1 mg/ml, where less than 50% of the high affinity (viteline layer) binding sites are occupied, and there is little binding to the low affinity (plasma membrane) sites. Thus, the interaction of sperm with vitelline layer components may be an obligatory step in the fertilization process. A fraction of the 125I-Con A binding sites is cleaved from the egg surface upon fertilization or after activation by the calcium ionophore A23187. This release of Con A binding sites occurs during the limited proteolysis of surface components that accompanies the cortical reaction, and does not occur with ammonia activation of the egg, where the cortical reaction does not occur. Thus, the changes in Con A binding at fertilization are caused by the massive cortical granule exocytosis that occurs within minutes of sperm penetration, and not by activation of the egg per se.