Characterization of lectin receptors isolated from the outer acrosomal membrane of boar spermatozoa

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Abstract
The outer acrosomal membrane (OAM) of boar spermatozoa was isolated by homogenization and centrifugation through modified colloidal silica. Homogeneity of the isolated membrane fraction (OAM) was revealed by transmission electron microscopy. At least 10 protein components could be discriminated by SDS-PAGE electrophoresis of the OAM, with molecular weights ranging from 340 to 15 kdal. Radiolabelling of the externally disposed carbohydrate side-chains by [3H]borhydride reduction of the isolated membrane, oxidized by use of galactose oxidase, revealed one main galactoprotein with a reduced molecular weight of about 270 kdal. This was identified as the RCA-120 receptor protein by means of lectin-affinity chromatography, high resolution gelfiltration and SDS-PAGE. Screening of the Con A binding properties of the solubilized membrane components partially isolated by affinity chromatography and HPLC was performed by an enzyme-linked-lectin-assay (ELLA). Electrophoretic analysis including a Con A-peroxidase staining procedure allowed the identification of 4 Con A binding proteins of the OAM with molecular weights of 120, 110, 88 and 66 kdal.