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Human teratocarcinoma derived cells, line PA 1, were labeled with radioactive monosaccharides and subsequently digested with pronase. Large sized glycopeptides (fraction A) were isolated by gel filtration on Bio-Gel P-10. Their chromatography on concanavalin A-Sepharose gave three subfractions, two of which were eluted with a sugar-free buffer and the third with 10 mM alpha-methyl mannoside. The first subfraction (fraction A-Con A Ia) incorporated label from [3H]galactose and [3H]glucosamine and contained the largest components of fraction A. The second and the third subfractions (fractions A-Con A Ib and A-Con A II) were glycopeptides which incorporated label from tritiated fucose, mannose, galactose, and glucosamine. Even these molecules were of large size eluting partially at the void volume from Bio-Gel P-60. The glycopeptides of fraction A-Con A Ib contained mannose, fucose, galactose, N-acetylglucosamine, and N-acetylgalactosamine. Fucose and galactose residues occupied ultimate or penultimate positions at the nonreducing termini of the oligosaccharides. N-Acetylneuraminic acid, too, was present in the glycopeptides of fraction A.

Human teratocarcinoma derived cells, line PA 1, maintained in the undifferentiated state, yielded upon exhaustive pronase digestion unusually large glycopeptides (fraction A), which showed on gel filtration an apparent molecular weight larger than 7400. These glycopeptides derived from whole cell proteins carried large-sized oligosaccharides as evidenced by repeated pronase treatments, hydrazinolysis, and beta-elimination experiments. The oligosaccharides consisted of mannose, fucose, galactose, and N-acetylglucosamine. The PA 1 cells contained also oligomannosyl type glycans, presumably linked to asparagine (fraction C glycopeptides). These glycopeptides were strongly bound to Con A--Sepharose and their oligosaccharides were released by endo-beta-N-acetylglucosaminidase H. The liberated glycans ranged from Man5GlcNAc to Man9GlcNAc as analyzed by paper chromatography. "Pulse-chase" experiments suggest that there is a precursor-product relationship between the mannose label in the fraction C (oligomannosyl type) glycopeptides and the fraction A glycopeptides.

Simultaneous separation of N-acetylglucosaminitol, N-acetylgalactosaminitol, N-acetylglucosamine and N-acetylgalactosamine from each other was achieved by paper chromatography. The method, which is suitable for analysis of radioactively labeled glycopeptides, allowed identification of N-acetylglucosamine and N-acetylgalactosamine as the linkage sugars in specific glycopeptide fractions isolated from human teratocarcinoma cells of line PA 1.

N-acetyl-lactosamine type glycopeptides of Semliki Forest virus were fractionated on concanavalin A-Sepharose, and their oligosaccharides were liberated by hydrazinolysis and analyzed by paper chromatography. Comparison with labeled glycans from reference glycopeptides suggests that the viral N-acetyl-lactosamine type glycans are bi-, tri- and tetra-antennary.