Quantitative glycoproteomics reveals new classes of STT3A- and STT3B-dependent N-glycosylation sites

J Cell Biol. 2019 Aug 5;218(8):2782-2796. doi: 10.1083/jcb.201904004. Epub 2019 Jul 11.


Human cells express two oligosaccharyltransferase complexes (STT3A and STT3B) with partially overlapping functions. The STT3A complex interacts directly with the protein translocation channel to mediate cotranslational glycosylation, while the STT3B complex can catalyze posttranslocational glycosylation. We used a quantitative glycoproteomics procedure to compare glycosylation of roughly 1,000 acceptor sites in wild type and mutant cells. Analysis of site occupancy data disclosed several new classes of STT3A-dependent acceptor sites including those with suboptimal flanking sequences and sites located within cysteine-rich protein domains. Acceptor sites located in short loops of multi-spanning membrane proteins represent a new class of STT3B-dependent site. Remarkably, the lumenal ER chaperone GRP94 was hyperglycosylated in STT3A-deficient cells, bearing glycans on five silent sites in addition to the normal glycosylation site. GRP94 was also hyperglycosylated in wild-type cells treated with ER stress inducers including thapsigargin, dithiothreitol, and NGI-1.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Glycoproteins / metabolism*
  • Glycosylation
  • HEK293 Cells
  • HSP70 Heat-Shock Proteins / metabolism
  • HeLa Cells
  • Hexosyltransferases / metabolism*
  • Humans
  • Membrane Proteins / metabolism*
  • Proteomics*


  • Glycoproteins
  • HSP70 Heat-Shock Proteins
  • Membrane Proteins
  • glucose-regulated proteins
  • Hexosyltransferases
  • STT3A protein, human
  • STT3B protein, human