Ribophorin I acts as a substrate-specific facilitator of N-glycosylation.

The mammalian oligosaccharyltransferase (OST) complex is composed of about eight subunits and mediates the N-glycosylation of nascent polypeptide chains entering the endoplasmic reticulum (ER). The conserved STT3 subunit of eukaryotic OST complexes has been identified as its catalytic centre, yet although many other subunits are equally well conserved their functions are unknown. We used RNA interference to investigate the function of ribophorin I, an ER-translocon-associated subunit of the OST complex previously shown to associate with newly synthesised membrane proteins. We show that ribophorin I dramatically enhances the N-glycosylation of selected membrane proteins and provide evidence that it is not essential for N-glycosylation per se. Parallel studies confirm that STT3 is essential for transferase activity of the complex, but reveal that the two mammalian isoforms are not functionally equivalent when modifying bona fide polypeptide substrates. We propose a new model for OST function where ribophorin I acts as a chaperone or escort to promote the N-glycosylation of selected substrates by the catalytic STT3 subunits.