Human cleavage signal-1 protein; cDNA cloning, transcription and immunological analysis.
The cleavage signal-1 protein (CS-1), a doublet antigen comprised of approx. 14-kDa and 18-kDa proteins has been shown to be present on the surface of sperm of various mammalian species including humans. Polyclonal antibodies to CS-1 inhibit the early cleavage of fertilized eggs without apparently affecting sperm penetration and pronuclear formation. We report here the cloning of the human CS-1 cDNA and its expression in vitro to obtain the recombinant protein (reCS-1) molecule. The CS-1 cDNA clone was isolated by immunological screening of a human testis lambda gt11 cDNA library with mono-specific polyclonal antibody against CS-1. The cDNA is 1828 bp long; the start codon assigned to the first ATG (bp 98-100) encodes a protein with 249 amino acid residues terminating at TAA (bp 845-847). The cDNA isolated has a 97-bp 5' and a 984-bp 3' untranslated region. The potential polyadenylation signal (5'-AATAAA) is at bp 1803-1808. An extensive computer search of the GenBank database did not indicate any extensive homology with any known sequence, indicating that CS-1 is a unique protein. The CS-1 cDNA was cloned in the transcription vector, pGEM-11Zf, to obtain high-level in vitro transcription by SP6 and T7 RNA polymerase. The transcribed CS-1 RNA was translated in a rabbit reticulocyte in vitro translation system and produced a 33-kDa reCS-1 protein, as assessed by migration in a SDS-polyacrylamide gel.(ABSTRACT TRUNCATED AT 250 WORDS)
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Department of Obstetrics and Gynecology, Albert Einstein College of Medicine, Bronx, NY 10461.
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D000595:Amino Acid Sequence; D000954:Antigens, Surface / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D001483:Base Sequence; D015153:Blotting, Western; D003001:Cloning, Molecular; D006801:Humans; D008297:Male; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D008840:Microfilament Proteins; D008969:Molecular Sequence Data; D015183:Restriction Mapping; D013094:Spermatozoa / Q000276:immunology*
D000954:Antigens, Surface; C482653:ITPRID2 protein, human; D008565:Membrane Proteins; D008840:Microfilament Proteins