Characterization of the chondroitin sulfate produced by B16 mouse melanoma cells

Abstract

The mucopolysaccharides produced by B16 mouse melanoma cells have been isolated in milligram quantities from the spent media in which the cells were grown in the presence of 2-amino-2-deoxy-D-glucose-t and [35S]-sulfate. The mucopolysaccharides obtained by precipitation with cetylpyridinium chloride from the Pronase digest of the media were further purified by gel filtration, ion-exchange chromatography, and treatment with nucleases. The major components were identified as chondroitin-4-sulfates by identification of the hexosamine as 2-amino-2-deoxy-D-galactose, and by digestibility with hyaluronidases, chondroitinase AC, and chondro-4-sulfatase. The o.r.d. curve and i.r. spectra of these components also confirmed their similarity to chondroitin-4-sulfate from cartilage. The molecular weight of the polysaccharide chains was estimated to be in the range 90,000-120,000 by sedimentation equilibrium analysis.