The critical glycosylation site of human transferrin receptor contains a high-mannose oligosaccharide

Glycobiology. 1995 Mar;5(2):227-32. doi: 10.1093/glycob/5.2.227.


The human transferrin receptor (TfR) contains three N-linked oligosaccharides and glycosylation is required for the proper folding and function of the molecule. Earlier studies demonstrated that the oligosaccharide at Asn-727 is vital for the production of fully active TfR. The oligosaccharide(s) present at this site have been analysed using a combination of site-directed mutagenesis and chemical analysis. Wild-type TfR and mutants containing only the Asn-727 site or missing all three sites were transfected into mouse 3T3 cells and receptors were analysed by endo-N-acetylglucosaminidase H (Endo-H) digestion, SDS-PAGE and immunoblotting. These studies suggested that the Asn-727 site contains high-mannose or Endo-H-sensitive hybrid oligosaccharides. Glycosylation of Asn-727 found in the TfR purified from human placentae was analysed by high-pH anion-exchange chromatography with pulsed amperometric detection (HPAE-PAD) and mass spectrometry following tryptic digestion, peptide purification via reverse-phase high-performance liquid chromatography (RP-HPLC) and peptide sequencing. HPAE-PAD showed the presence of a series of high-mannose oligosaccharides. Mass spectrometry confirmed these observations, but also showed the presence of an 80 Da anionic moiety on a fraction of the oligosaccharides.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • 3T3 Cells
  • Amino Acid Sequence
  • Animals
  • Asparagine / analysis
  • Binding Sites
  • Carbohydrate Sequence
  • Chromatography, High Pressure Liquid
  • Electrophoresis, Polyacrylamide Gel
  • Glycosylation
  • Hexosaminidases
  • Humans
  • Mannose / analysis*
  • Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase
  • Mass Spectrometry
  • Mice
  • Molecular Sequence Data
  • Oligosaccharides / chemistry*
  • Oligosaccharides / isolation & purification
  • Peptide Mapping
  • Receptors, Transferrin / biosynthesis
  • Receptors, Transferrin / chemistry*
  • Receptors, Transferrin / metabolism*
  • Recombinant Proteins / biosynthesis
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / metabolism
  • Transfection
  • Trypsin


  • Oligosaccharides
  • Receptors, Transferrin
  • Recombinant Proteins
  • Asparagine
  • Hexosaminidases
  • Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase
  • Trypsin
  • Mannose