The goal of this research program is to clone and characterize proteins which are located on the surface of the human sperm and to evaluate their immunocontraceptive potential. Attention will be directed to proteins which are auto- or iso-antigens and are recognized by sperm agglutinating antibodies. A computerized 2-D database based on migration of human sperm proteins in two dimensional gels has been developed. Within this database a subset of 98 human sperm surface proteins accessible to surface iodination and biotinylation has been identified. The locations of these surface proteins in two dimensional gels have been catalogued in a sperm protein encyclopedia. A subset of these sperm surface proteins will be microsequenced, and complete open reading frames will be obtained by molecular cloning. Unique sperm proteins will be characterized and evaluated as potential contraceptive immunogens according to strict criteria which include testis specificity and involvement in functions mediated at the sperm surface.
The specific aims are: 1) To microsequence human sperm surface proteins through a combination of Edman degradation and/or tandem mass spectrometry; 2) To clone and sequence the complete open reading frames for an estimated 20 cDNAs representing unique sperm surface proteins using a combination of molecular approaches; 3) To determine tissue specificity of the cloned cDNAs by Northern blot, RT-PCR, and immunohistochemical analyses on an organ bank of human and non- human primate mRNAs and tissues; 4) To confirm the sperm surface localization of candidate immunogens and test their contraceptive potential through in vitro functional assays; and 5) To conduct immunogenicity and fertility trials of several novel sperm surface proteins in mice and macaques. Achievement of these aims will increase our understanding of the molecular character of proteins exposed on the exterior of the human sperm surface and aid in selection of candidate antigens appropriate for human immunocontraception by immortalizing and manipulating the cDNAs encoding these proteins.
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