Question

Temperature-sensitive yeast mutants have been isolated that block each of the enzymatic steps in the synthesis of the dolichol-linked oligosaccharide precursor for n-linked glycosylation. why do mutations that block synthesis of the intermediate with the structure dolichol-pp-(glcnac)2man5 completely prevent addition of n-linked oligosaccharide chains to secretory proteins, whereas mutations that block conversion of this intermediate into the completed precursor—dolichol-pp-(glcnac)2man9glc3—allow the addition of n-linked oligosaccharide chains to secretory glycoproteins?

Answer 1

This is because the seven-sugar intermediate is synthesized by sugar addition to cytosolic-facing dolichol phosphate. The intermediate is flipped from the cytosol face of the ER membrane to the the luminal face. Additionally, the sugar additions then occur within the lumen of the ER. The short forms of the intermediate are on the wrong side of the membrane to add to nascent polypeptides within the ER lumen. Incomplete adductants within the ER lumen are located appropriately to N-glycosylate nascent polypeptide.