Title : Presence of O-linked oligosaccharide on a threonine residue in the human transferrin receptor
Abstract :
We have previously demonstrated that the human transferrin receptor ( TfR ) of approximately 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 beta-elimination of the intact TfR , but not the 70 kDa fragment , released Gal-[3H]Gal-NAcitol, 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 beta-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 beta-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