Cloning and expression of recombinant rabbit fertilin.
Fertilin is a sperm surface protein complex which is reported to play an essential role in sperm-egg fusion in mammals. It is comprised of two related subunits, alpha and beta, both of which are glycosylated and have cytoplasmic and extracellular domains. This protein has been reported to play an essential role in sperm-egg fusion in mammals. We report on the cloning and sequencing of the complete cDNA sequences of both subunits from rabbit testis, and the production of recombinant proteins for testing their potential as antigens for use in an immunocontraceptive vaccine to control wild rabbit populations. The cDNAs for rabbit fertilin alpha and beta (Genbank accession numbers, U46069 and U46070) are predicted to encode proteins of 919 and 751 amino acids, respectively, and to show significant levels of homology to fertilin subunits isolated from other species. Analysis of the predicted protein sequences of fertilin alpha but not beta reveals the presence of 21 direct repeats of the hexameric sequence A/PPPPEA at the extreme carboxy terminus, similar to what has been described for a fertilin alpha gene isoform in the monkey. DNA sequences corresponding to the predicted mature alpha and beta fertilin subunits were individually cloned into a bacterial expression system, and the recombinant proteins were used to raise polyclonal antibodies in mice. These antibodies detect components of the native fertilin complex from rabbit sperm.
Co-operative Research Centre for the Biological Control of Vertebrate Pest Populations, Commonwealth Scientific and Industrial Research Organisation, Lyneham, Canberra, Australia.
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Holland
D051722:ADAM Proteins; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics / Q000302:isolation & purification; D004926:Escherichia coli / Q000235:genetics; D000072196:Fertilins; D008562:Membrane Glycoproteins / Q000096:biosynthesis / Q000235:genetics*; D008666:Metalloendopeptidases / Q000096:biosynthesis / Q000235:genetics*; D051379:Mice; D008969:Molecular Sequence Data; D011817:Rabbits; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D016415:Sequence Alignment
D018076:DNA, Complementary; D008562:Membrane Glycoproteins; D011994:Recombinant Proteins; D051722:ADAM Proteins; C000606455:Adam1a protein, mouse; D000072196:Fertilins; D008666:Metalloendopeptidases