title: Biochemical and binding characteristics of boar epididymal fluid proteins.
abstract: During the passage through the epididymis, testicular spermatozoa are directly exposed to epididymal fluid and undergo maturation. Proteins and glycoproteins of epididymal fluid may be adsorbed on the sperm surface and participate in the sperm maturation process, potentially in sperm capacitation, gamete recognition, binding and fusion. In present study, we separated proteins from boar epididymal fluid and tested their binding abilities. Boar epididymal fluid proteins were separated by size exclusion chromatography and by high-performance liquid chromatography with reverse phase (RP HPLC). The protein fractions were characterized by SDS-electrophoresis and the electrophoretic separated proteins after transfer to nitrocellulose membranes were tested for the interaction with biotin-labeled ligands: glycoproteins of zona pellucida (ZP), hyaluronic acid and heparin. Simultaneously, changes in the interaction of epididymal spermatozoa with biotin-labeled ligands after pre-incubation with epididymal fluid fractions were studied on microtiter plates by the ELBA (enzyme-linked binding assay) test. The affinity of some low-molecular-mass epididymal proteins (12-17 kDa and 23 kDa) to heparin and hyaluronic acid suggests their binding ability to oviductal proteoglycans of the porcine oviduct and a possible role during sperm capacitation. Epididymal proteins of 12-18 kDa interacted with ZP glycoproteins. One of them was identified as Crisp3-like protein. The method using microtiter plates showed the ability of epididymal fluid fractions to change the interaction of the epididymal sperm surface with biotin-labeled ligands (ZP glycoproteins, hyaluronic acid and heparin). These findings indicate that some epididymal fluid proteins are bound to the sperm surface during epididymal maturation and might play a role in the sperm capacitation or the sperm-zona pellucida binding.
authors.affiliation: Laboratory of Diagnostics for Reproductive Medicine, Institute of Biotechnology, Academy of Sciences of the Czech Republic, v.v.i., Vídeňská 1083, 142 20 Prague 4, Czech Republic. pavlam@img.cas.cz
authors.forename: Pavla
authors.initials: P
authors.lastname: Maňásková-Postlerová
authors.forename: Nina
authors.initials: N
authors.lastname: Davidová
authors.forename: Věra
authors.initials: V
authors.lastname: Jonáková
mesh_terms: D000818:Animals; D001710:Biotin; D026481:Blotting, Far-Western; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D056148:Chromatography, Reverse-Phase; D004527:Egg Proteins / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D057075:Enzyme Assays; D030501:Epididymal Secretory Proteins / Q000737:chemistry* / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000378:metabolism; D008970:Molecular Weight; D011485:Protein Binding; D011956:Receptors, Cell Surface / Q000378:metabolism; D034421:Sus scrofa; D000072216:Zona Pellucida Glycoproteins
chemical_list: D004527:Egg Proteins; D030501:Epididymal Secretory Proteins; D008562:Membrane Glycoproteins; D011956:Receptors, Cell Surface; D000072216:Zona Pellucida Glycoproteins; D001710:Biotin