Comparison of the N-linked glycopeptides of DQw1 and DR1 molecules.

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HLA class II molecules have been isolated from a [3H]mannose-labeled GM3104 B lymphoblastoid cell line with the phenotype DQw1, DR1. The DQw1 molecules were purified by affinity to 77-34 IgG specifically reactive with the DQw1 specificity. The DR1 molecules were separated into two subsets, DR1a (70 to 80%) and DR1b (20 to 30%), by sequential affinity to 21r5-IgG and 21w4-IgG Sepharose. The alpha- and beta-chains of [3H]mannose-labeled DQw1, DR1a, and DR1b molecules were separated by SDS-PAGE and were recovered by electrophoretic elution. The isolated chains were digested with pronase and the glycopeptides were fractionated by sequential lectin chromatography on immobilized concanavalin A (Con A), Lens culinaris (Lens), and Ricinus communis agglutinin type I (RCA). The N-linked glycopeptides derived from the alpha-chains of DQw1, DR1a, or DR1b showed similar profiles on Con A Sepharose: 45% unbound (ConA I), 25% weakly bound (ConA II), and 30% tightly bound (ConA III). The glycopeptides derived from the beta-chains of DQw1 or DR1 molecules were found almost exclusively (80%) in the fraction unbound to Con A Sepharose, with only 11% and 9% in ConA II and ConA III fractions, respectively. The observation that most of the binding to Con A is associated with the alpha-chain glycopeptides suggests that binding of membrane-associated class II molecules to that lectin must be mediated by the alpha-chains. Binding to Lens Sepharose was higher for beta-(50%) than for alpha-(15%) chain glycopeptides, suggesting that within the intact glycoproteins, the beta-chains are responsible for the interaction with Lens. The ConA I fractions derived from the alpha-chain glycopeptides of either DQw1 or DR1 molecules were separated on RCA-agarose as follows: 60% unbound, 17% retarded, and 20% bound and eluted with 0.1 M galactose. The ConA I fractions derived from the beta-chain glycopeptides of either subset of class II molecules also had a similar profile on RCA-agarose: 70% unbound, 16% retarded, and 10% bound and eluted specifically. After removal of sialic acid residues, all of the ConA I fractions of alpha- and beta-chains bound to RCA-agarose. A high degree of similarity was observed between the corresponding glycopeptides of the three subsets of class II molecules and between the complex N-linked structures of alpha- and beta-chains. Minor variations were observed between DR1a and DR1b glycopeptides which appear greater than those observed between DR1 and DQw1 glycopeptides.(ABSTRACT TRUNCATED AT 400 WORDS)