PLASMA MEMBRANE CHANGES IN FERTILIZATION

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These studies focus on the structure and function of a previously identified 350kDa glycoprotein on the surface of the sea urchin egg that mediates binding of sperm. Since this binding event is a pre-requisite to fertilization, this information in the long term may be of value in the development of new methods to enhance or prevent fertilization in higher organisms, including humans. Two major questions about the receptor will be addressed. The first question pertains to its mode of localization and biosynthesis. To elucidate the molecular basis for association of the receptor with the cell surface of S. purpuratus eggs, several analytical and biochemical techniques will be applied. These biochemical studies will be complemented by in vivo studies on the biosynthesis of the receptor in starfish oocytes to determine if the full length sea urchin receptor can be synthesized, co- and post- translationally modified, and routed to the cell surface of these oocytes in a functional form. The second question pertains to the mechanism of sperm binding to the receptor. The structure and the function of three structural elements of the receptor known to be involved in sperm binding will be studied. With respect to two of the element, polypeptide domains, deletion and mutational analysis will be performed to define the minimum size and sequence of each required for sperm binding. Also, the significance of the hsp110 homology of the receptor with respect to its function in binding sperm will be studied, particularly with respect to a protein in rat sperm that is believed to be immunologically cross-reactive with hsp70. It will be determined if the third structural element of the receptor, the sulfated O-linked oligosaccharide chains, compete with the two polypeptide domains for binding to sperm, or interact with a different binding site on the sperm. A full understanding of the control of specificity of sperm binding requires a comparison with the primary and complete structure of the receptor from another species of sea urchin. This will be carried out in L. pictus, a sea urchin of another genus that does not cross fertilize with S. purpuratus.