Plasma membranes were isolated from an
ascites hepatoma, AH 130 FN, a free-cell type subline of AH 130, by the
fluorescein mercuric acetate (FMA) method.
Glycopeptides and
mucopolysaccharides were prepared from the membranes by
pronase digestion then fractionated chromatographically and electrophoretically. Isolated fractions were analyzed for
amino acid and
carbohydrate compositions. The results were compared with those for corresponding fractions from AH 66 and AH 130 ((1974) J. Biochem. 76, 319-333; (1975) ibid., 78, 863-872). The fraction excluded from
Sephadex G-50 contained
mucopolysaccharides and a series of
glycopeptides. The
mucopolysaccharides were identified as
chondroitin sulfate A on the basis of their chemical composition, electrophoretic behavior on
cellulose acetate and digestibility with
chondroitinase AC [EC 4.2.2.5]. This contrasts with previous findings that
mucopolysaccharides from the corresponding fractions from AH 130 and AH 66 were
heparan sulfate. The chemical composition of the
glycopeptides, which showed high contents of
threonine,
serine,
galactose,
galactosamine,
glucosamine, and
sialic acid, indicated the presence of
glycopeptides with O-glycosidic linkages. The
glycopeptides also contained a small but significant amount of
aspartic acid, suggesting that N-glycosidic
glycopeptides were also contained in this fraction. The fraction included in Sepnadex G-50 contaoned N-glycosidic
glycopeptides as major components, since the
carbohydrate moieties were composed of
fucose,
galactose,
mannose,
glucosamine,
sialic acid, and a smaller amount of
galactosamine. The presence of
galactosamine suggested that O-glycosidic
glycopeptides were present as minor components.
Glycopeptides with both O- and N-glycosidic linkages were isolated from AH 130, but not from AH 66.