ANALYSIS OF THE GLYCOSYLATION PATTERNS OF THE EXTRACELLULAR DOMAIN OFTHE EPIDERMAL GROWTH-FACTOR RECEPTOR EXPRESSED IN CHINESE-HAMSTER OVARY FIBROBLASTS

The extracellular domain (621 N-terminal amino acids) of the p170 epid ermal growth factor (EGF) receptor has eleven consensus N-linked glyco sylation sites. When expressed in Chinese hamster ovary cells this was glycosylated with a combination of high mannose and complex chains. T he latter chains were shown by chromatographic separation and mass spe ctrometric analysis of tryptic digests to be clustered in the EGF-bind ing domain. Treatment with the endoglycosidase, peptide-N-glycosidase F (PNGase F), reduced the molecular weight from 110 kDa to 75 kDa. Rel eased oligosaccharides were characterised at high sensitivity by high pH anion exchange chromatography with pulsed amperometric detection an d gas liquid chromatography/mass spectrometry. The data were consisten t with the complex chains being trisialylated tetra-antennary oligosac charides fucosylated on the reducing terminal GlcNAc. The large hydrod ynamic mass of these oligosaccharides could influence ligand binding, an effect which is likely to vary with the difference in consensus gly cosylation sites of proteins related to p170 i.e. p185(erbB2/neu), p18 0(erbB3) and p180(erbB4).