Purification of the major sialoglycoproteins of 13762 MAT-B1 and MAT-C1 rat ascites mammary adenocarcinoma cells by density gradient centrifugation in cesium chloride and guanidine hydrochloride.

Abstract

Both MAT-B1 (nontransplantable) and MAT-Cl (xenotransplantable) ascites sublines of the 13762 rat mammary adenocarcinoma contain a major sialoglycoprotein (ASGP-1) with low electrophoretic mobility in dodecyl sulfate.ASGP-1 from each subline is rapidly purified by density gradient centrifugation of membrane fragments in cesium chloride containing 4 M guanidine hydrochloride.The products give single bands on dodecyl sulfate-polyacrylamide gel electrophoresis and single peaks by gel filtration in dodecyl sulfate o r guanidine hydrochloride.The composition of ASGP-1 from each subline is high in serine, threonine, galactosamine, glucosamine, galactose, and sialic acid, with carbohydrate compositions of 67 and 73% for MAT-B1 and MAT-C1 ASGP-1, respectively.Amino acid compositions of ASGP-1 from the sublines are essentially the same.The major difference between them is a %fold greater content of sialic acid/unit of protein for MAT-Cl ASGP-I than MAT-B1 ASGP-1.Molecular weights of 670,000 (MAT-B1) and 690,000 (MAT-C1) are estimated by sedimentation velocity analysis and gel filtration in 4 M guanidine hydrochloride.Fractionation of oligosaccharides from alkaline borohydride-treated ASGP-1 on Bio-Gel P-4 gives patterns that are quantitatively different for MAT-B1 and MAT-C1.The larger of the two major oligosaccharides from MAT-Cl (IIC) is larger than either of the two major oligosaccharides from MAT-B1, while the smaller of the two major MAT-B1 oligosaccharides (IVB) is smaller than either of the major MAT41 oligosaccharides.The size differences are consistent with the oligosaccharide compositions, showing increased amounts of sialic acid for the larger oligosaccharides.Differences in MAT-B1 and MAT-C1 ASGP-1 at the oligosaccharide level may account for the differences in ability of the sublines to be transplanted across species histocompatibility barriers.It has been suggested that cell surface glycoproteins may