Localization of the Carbohydrate Units in a Human Immunoglobulin Light Chain, Protein Sm λ

Abstract

The carbohydrate structure and complete amino acid sequence of a human lambda-type immunoglobulin light chain, protein Sin λ, has been determined. The protein was isolated from the urine of a patient with a plasma cell dyscrasia resembling γ-heavy-chain disease. 13 tryptic peptides covering the entire polypeptide chain of 135 residues were isolated from the aminoethylated protein, and 15 chymotryptic peptides, accounting for 131 residues, were recovered from the carboxymethylated protein. The sequence of 18 of these peptides was partially or completely determined by the Edman-dansyl technique or C-terminal analysis, permitting the establishment of the complete primary structure of the polypeptide chain. The sequence established that this light chain possessed an intramolecular deletion of 81 amino acid residues. The N-terminal 30 residues showed considerable homology with other lambda chains of subgroup 11. The defect began at position 31, in the first hypervariable region, and encompassed the remainder of the variable region through position 109. The constant region was fully intact and normal synthesis recommenced with a glutaminyl residue at position 110, the first residue of the constant region. This light chain contained carbohydrate in the hypervariable region just preceding the deletion. The precise number and locations of the oligosaccharide chains were established by amino acid sequence analysis of glycopeptides isolated from proteolytic hydrolysates by chromatography on Bio-Gel P-6 columns. These studies showed that protein Sm λ contains one N-glycosidically-linked chain attached to asparagine-25 and one O-glycosidically-linked oligosaccharide chain attached to serine-21. The structures of the oligosaccharide chains were determined by methylation analysis, gas chromatography and hydrolysis with specific glycosidases The structure of the N-glycosidically-linked chain was NeuAc(α2→6)Gal(β1→4)G1cNAc(β1→2)Man(α1→6) [NeuAc(α2→6)Gal(β1→4)GlcNAc(β→2)Man(α1→3)]Man(β1→4)GlcNAc(β→4) [Fucα 1→6]GlcNAc→Asn. The second O-glycosidically-linked chain was a disialylated tetrasaccharide with the structure, Neu(α2→3)Gal(β1→3)[NeuAc(α2→6)GalNAc→Ser. This mucin-type disialylated tetrasaccharide in close proximity to N-asparagine-linked chains has not been previously observed in the oligosaccharide chains of immunoglobulins.