Maternal recognition of pregnancy in sheep is dependent upon secretion of ovine trophoblast protein-1 (oTP-1) by the conceptus. Between days 13 and 21 of pregnancy, oTP-1 represents the major conceptus secretory product, and it is responsible for inhibition of uterine prostaglandin F2alpha secretion. The oTP-1-induced inhibition of pulsatile secretion of prostaglandin F2alpha by the uterus allows for maintenance of the corpus luteum with continued secretion of progesterone. Porcine and bovine conceptuses have also been shown to produce similar antiluteolytic agents, and a human equivalent has been postulated as well. oTP-1 has recently been shown to share amino acid homology with alpha interferons. Thus, oTP-1 is involved in the interaction between the conceptus and maternal systems which results in appropriate endocrinological and immunological responses in early pregnancy. We propose to examine the structure/function basis for pregnancy recognition and other biological effects of oTP-1 using the synthetic peptide approach for both direct receptor competition and production of monoclonal antibodies to oTP-1 and oTP-1 synthetic peptides. It is planned that the objective be achieved through the following approach: (1) perform competitive receptor binding and functional experiments between synthetic peptides and oTP-1, and determine the ability of appropriate peptides to bind to the oTP-1 receptors; particular emphasis will be placed on the use of longer peptides (possible domains); (2) modify peptides that are important in receptor competition by systematic removal and/or substitution of amino acids in order to more precisely identify sequences or regions of oTP-1 that are involved in functions: (3) generate monoclonal antibodies to oTP-1 and determine their ability to block function and/or binding of oTP-1 to membrane receptors; (4) map the epitope specificity of these monoclonal antibodies using synthetic peptides that correspond to regions of the oTP-1 molecule located on the surface; (5) use synthetic peptides to produce site specific monoclonal and polyclonal antibodies and determine their effect on functions and receptor binding by oTP-1: (6) determine the steric relationship of epitopes, (and thus various regions) of oTP-1 by competitive binding between monclonal antibodies (and their Fab fragments) of defined epitope specificities with oTP-1. The proposed studies are important because they will provide information on the structural basis for pregnancy recognition and the antiviral properties of oTP-1.
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