The Structure of the Glycopeptide of Human γG Myeloma Proteins

Abstract

Abstract Glycopeptides were isolated from seven γG myeloma proteins and the γG-immunoglobulins of one normal individual. The composition of each glycopeptide was determined and the structure of the oligosaccharide portion of the molecule established as follows. Sequential enzymatic degradation of the oligosaccharide chains with purified glycosidases established the sequences of the sugars. Serial periodate oxidation and methylation of both intact and partially degraded glycopeptides followed by identification of the methylated sugars by gas chromatography and mass spectrometry established the linkages between sugars. The carbohydrate composition of the glycopeptides was found to be (in residues per mole of glycopeptide): sialic acid, 0 to 2; fucose, 0.6 to 1.2; galactose, 0.3 to 2.3; N-acetylglucosamine, 3.9 to 6.0; and mannose, 3. The variability in the number of sialic acid, fucose, and galactose residues was shown to be caused by microheterogeneity of the oligosaccharides while the variability in the number of N-acetylglucosamine residues reflects differences in the sequence of the sugars in the of the molecules. Two of the seven myeloma proteins had an additional oligosaccharide chain. All the glycopeptides are branched oligosaccharides containing two nonreducing termini with the sequence Gal (β1,6)/→ GlcNAc (β1,2)/→ Man. When sialic acid is present, it is linked α2,6 to the galactose residue. Both branches are connected to another mannose residue in the core, which also contains either 2 or 3 N-acetylglucosamine (GlcNAc) residues. The sequence of sugars in the core varied from one glycopeptide to another, being either Man→GlcNAc→ GlcNAc→Asn, Man→[GlcNAc→]GlcNAc→Asn, or Man→ GlcNAc→[GlcNAc→]GlcNAc→Asn. Di-N-acetylchitobiose was isolated from the core of one glycopeptide with a core structure of Man→GlcNAc→GlcNAc→Asn. These studies establish that homogeneous γG-immunoglobulins of man may have different sequences of sugars in their oligosaccharide chains. The data also show that microheterogeneity of oligosaccharides exists in homogeneous glycoproteins secreted by a single clone of cells.