The sialoglycoprotein subunits of human placental brush border membranes characterized by two-two-dimensional electrophoresis.
A brush border membrane enriched fraction was isolated from human full-term placenta. This membrane fraction exhibited large membrane fragments with microvilli projecting from the basal membrane in electron micrographs and was enriched tenfold in alkaline phosphatase, a brush border enzyme marker. The sialoglycoproteins associated with this membrane fraction were tritiated by mild periodate oxidation of sialic acid and reduction with tritiated NaBH4. The membranes were solubilized in 8 M urea, 2 percent Triton X-100, and the tritiated glycoprotein subunits were reduced with beta-mercaptoethanol and characterized by 2-dimensional poly-acrylamide gel electrophoresis using a method similar to that described by O'Farrell and Bhakdi, Knüferman, and Wallach. The tritiated subunits were detected in the gels by autofluorography. The 2-dimensional subunit "maps" resolved at least 17 major sialoglycoprotein subunits whereas only 10 major periodate-Schiff reagent staining components were resolved by 1-dimensional SDS polyacrylamide gel electrophoresis. Placental alkaline phosphatase (PAP) was identified on the subunit maps by inclusion of 32P-labeled PAP in the tritiated membrane sample. The 32P-labeled PAP corresponded to a major tritiated sialoglycoprotein subunit, which was heterogeneous with respect to charge as demonstrated by 3 closely running spots of the same molecular weight.
D002458:Cell Fractionation; D002462:Cell Membrane / Q000032:analysis*; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D006801:Humans; D046911:Macromolecular Substances; D008854:Microscopy, Electron; D008871:Microvilli / Q000032:analysis* / Q000648:ultrastructure; D010920:Placenta / Q000032:analysis*; D011247:Pregnancy; D012795:Sialoglycoproteins / Q000032:analysis*
D046911:Macromolecular Substances; D012795:Sialoglycoproteins