Glycobiology vol 2 no 4 pp. 345-353, 1992
© 1992
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Presence of O-linked oligosaccharide on a threonine residue in the human transferrin receptor
Department of Biochemistiy, University of Georgia Athens, GA 30602, USA
1To whom correspondence should be addressed
Received on March 29, 1992; accepted on May 13, 1992
We have previously demonstrated that the human transferrin receptor (TfR) of {small tilde}90 kDa contains Ser/Thr-linked (O-linked) oligosaccharides. In the present study, we report our identification of the site of attachment of the O-linked oligosaccharides in the receptor. A 70 kDa fragment from the external domain of the TfR was generated by trypsin treatment of the [3H]glucosamine-labelled receptor purified from human K562 cells. The ß-elimination of the intact TfR but not the 70 kDa fragment, released Gal-[3H]GalNAcitol, indicating that the 70 kDa fragment lacks O-linked oligosaccharides. In the remaining 20 kDa fragment there are three potential sites (Thr96, Thr104 and Ser106, for O-glycosylation in the extracellular domain. To identify which of these residues are O-glycosylated, both the [3H]Thr- and [3H]Ser-labelled TfR were directly treated with mild base to effect ß-elimination, and the radiolabelled amino acids and their derivatives were analysed. Approximately 2% of the total radiolabelled Thr, but no radiolabelled Ser, was converted to expected ß-elimination products by this treatment. These and other results demonstrate that only one O-linked oligosaccharide is present in the TfR and that it occurs on either Thr96 or Thr104 From human serum we purified the cleaved, soluble form of the TfR (s–TfR), which contains Thr104 but lacks Thr96 The s–TfR was sensitive to O-glycanase and bound to Jacalin lectin, indicating that the s–TfR contains an O-linked oligosaccharide. These results demonstrate that the human TfR, which contains a total of 40 Thr and 48 Ser residues in the extracellular domain, has a single O-linked oligosaccharide on Thr104 near the transmembrane domain.
O-glycosylation site transferrin receptor
2Current address: Oklahoma Center for Molecular Medicine, Department of Biochemistry and Molecular Biology, University of Oklahoma Health Science Center, Oklahoma City, OK 73104, USA
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