Characterization of a fucose-binding protein from bull sperm and seminal plasma that may be responsible for formation of the oviductal sperm reservoir.
Oviductal sperm reservoirs have been found in cattle, mice, hamsters, pigs, and horses. In cattle (Bos taurus), the reservoir is evidently formed when sperm bind to fucosylated ligands resembling Le(a) trisaccharide on the surface of oviductal epithelium. The aim of this study was to characterize the fucose-binding protein on bull sperm. Fresh ejaculated sperm were extracted with 0.5 M KCl in Hepes-balanced salts. Extracts were subjected to affinity chromatography using immobilized Le(a) trisaccharide (alpha-L-Fuc[1,4]-beta-D-Gal[1,3]-D-GlcNAc). Two-dimensional PAGE of the affinity chromatography eluates revealed a prominent protein of approximately 16.5 kDa and a pI of 5.8. This protein inhibited binding of sperm to oviductal explants. A similar analysis of proteins extracted from capacitated sperm (which do not bind to oviductal epithelium) showed a reduction in the amount of the 16.5-kDa protein. When examined by epifluorescence microscopy, live uncapacitated sperm labeled over the acrosome with a fucose-BSA-fluorescein isothiocyanate (FITC) conjugate, while capacitated sperm did not. When capacitated sperm were treated with 16.5-kDa protein, labeling with fucose-BSA-FITC was partially restored. The comparative ease with which the protein was removed from sperm and its apparent reassociation with sperm suggested that it could be a peripheral protein derived from epididymal or accessory gland fluids. Blots of SDS-PAGE gels of seminal plasma proteins revealed the presence of a Le(a)-binding protein with an apparent mass of 16.5 kDA: Amino acid sequencing of two tryptic fragments of the protein purified from sperm extracts identified it as PDC-109 (BSP-A1/A2), a product of the seminal vesicles.
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Department of Biomedical Sciences, College of Veterinary Medicine, Cornell University, Ithaca, New York 14853, USA.
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Perez
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Gwathmey
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Suarez
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D000177:Acrosome / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002846:Chromatography, Affinity; D015180:Electrophoresis, Gel, Two-Dimensional; D004848:Epithelium / Q000378:metabolism; D005187:Fallopian Tubes / Q000502:physiology*; D005260:Female; D016650:Fluorescein-5-isothiocyanate; D005456:Fluorescent Dyes; D005643:Fucose; D037102:Lectins / Q000032:analysis / Q000737:chemistry / Q000502:physiology*; D008297:Male; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D012661:Semen / Q000737:chemistry*; D012710:Serum Albumin, Bovine; D013075:Sperm Capacitation; D013094:Spermatozoa / Q000737:chemistry*
D005456:Fluorescent Dyes; D037102:Lectins; C027112:fucose-binding lectin; D012710:Serum Albumin, Bovine; D005643:Fucose; D016650:Fluorescein-5-isothiocyanate