Partial structure of a membrane glycopeptide from virus-transformed hamster cells

Abstract

The predominant surface glycopeptide from a clone of baby hamster kidney cells transformed by Rous sarcoma virus (C13/B4), metabolically labeled with L-[14C]fucose, was characterized for the 1st time. This glycopeptide represents 19% of the total radioactivity removed by trypsin from the cell surface of the transformed fibroblasts and is more abundant in the transformed cells than in the normal counterpart. Purification of the glycopeptide after digestion with Pronase was by successive chromatography on DEAE-cellulose and Sephadex G-50. The monosaccharide content of the glycopeptide was 42, 127, 138, 114 and 243 nmol of fucose, sialic acid, galactose, mannose and glucosamine, respectively. A partial structure of the glycopeptide was proposed from the results of sequential enzymatic degradation coupled with GLC anlaysis of the resultant monosaccharides. All of the enzymes used were purified and pretested on natural substrates and removed terminal monosaccharides of the correct configuration, quantitatively. The purification and properties of an .alpha.-L-fucosidase from rat testes were described. All of the radioactivity in the glycopeptide, recovered as fucose, was present at the core and was removed by treatment with this .alpha.-L-fucosidase. The proposed structure is a triantennary, completely sialylated, complex glycopeptide containing a core region of .beta.-D-mannose, .beta.-D-N-acetylglucosamine and .alpha.-L-fucose.