Structural analysis and localization of the carbohydrate moieties of a soluble human interferonγreceptor produced in baculovirus‐infected insect cells

Abstract

A soluble form of the human interferon γ receptor that is required for the identification of interferonγantagonists was expressed in baculovirus‐infected insect cells. The protein carried N‐linked carbohydrate and showed a heterogeneity on denaturing polyacrylamide gels. We investigated the utilization of the potential sites for N‐linked glycosylation and the structure of the carbohydrate moieties of this soluble receptor. Amino acid sequence analysis and ion spray mass spectrometry revealed that of the five potential sites for N‐linked glycosylation, Asn” and Asn69 were always utilized, whereas Asn62 and Asn162 were utilized in approximately one‐third of the protein population. Asn223 was never found to be glycosylated. The soluble receptor was treated with N‐glycosidase F and the oligosaccharides released were analyzed by matrix‐assisted laser desorption mass spectrometry, which showed that the protein carried six types of short carbohydrate chains. The predominant species was a hexasaccharide of molecular mass 1,039, containing a fucose subunit linked to the proximal N‐acetylglucosamine residue. Copyright © 1994 The Protein Society