One step purification and biochemical characterization of a spermatozoa-binding protein from porcine oviductal epithelial cells.
In pigs the binding of sperm to oviductal epithelial cells to form a sperm reservoir involves carbohydrate interactions. In the present study, we purify a sperm binding glycoprotein (SBG) from cells from the isthmus of the oviduct using an affinity column. This protein conjugated with FITC is able to bind to the heads of pig sperm. SBG is shown to contain carbohydrates by PAS-silver staining and lectin binding assays. Enzymatic treatment and lectin affinity demonstrate that SBG exposes Galbeta1-3GalNAc disaccharide, which is bound to a serine or a threonin residue by an O-link. After enzymatic deglycosylation SBG shows an apparent molecular mass of 67.5 kDa, which changes to 85 kDa by reduction with 2-mercaptoethanol. Both SBG and enzymatically deglycosylated SBG show by isoelectrofocusing two forms of pI 3.6 and pI 3.8. SBG may be involved on sperm-oviduct interaction.
División Biología del Desarrollo, Instituto de Biología Molecular y Celular de Rosario (IBR-CONICET), Consejo de Investigaciones de la Universidad Nacional de Rosario (CIUNR-UNR), Rosario, Argentina.
Patricia E
PE
Marini
Marcelo O
MO
Cabada
D000818:Animals; D050260:Carbohydrate Metabolism; D002352:Carrier Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D002846:Chromatography, Affinity; D004847:Epithelial Cells / Q000737:chemistry*; D005260:Female; D016650:Fluorescein-5-isothiocyanate; D006023:Glycoproteins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D008297:Male; D010057:Oviducts / Q000737:chemistry*; D013094:Spermatozoa / Q000378:metabolism*; D013552:Swine
D002352:Carrier Proteins; D006023:Glycoproteins; D016650:Fluorescein-5-isothiocyanate