Biosynthesis of N-linked glycoproteins

This process occurs in the ER and can be continued in the GA.

First a lipid-linked oligosaccharide precursor is synthesized. Its structure is the same in animals, plants and single-celled eucaryotes :

The precursor oligosaccharide is linked by a pyrophosphoryl group to dolichol. It is a long (75-95 carbon atoms), highly hydrophobic polyisoprenoid lipid :

Dolichol is long enough to span the membrane 4 - 5 times. It is localized on the rough ER. The biosynthesis starts at the cytosolic face of the ER. At the beginning 2 GlcNAc and 5 mannose residues are added one at time to dolichol phosphate. Then the dolichol pyrophosphoryl oligosaccharide is flipped to the luminal face. In the latter reactions each mannose or glucose residue is transferred from nucleotide sugar to dolichol on the cytosolic face of the ER. Then it is flipped and transferred to the growing oligosaccharide. The carrier is then flipped back again to the cytosolic face.
The oligosaccharide precursor is transferred from the dolichol to an Asn residue on the nascent polypeptide. Thus the glycosylation occurs cotranslationally. The Asn residue must be in a sequence N-X-S/T where X cannot be Pro or Asp.
The process is catalyzed by oligosaccharide protein transferase. The enzyme is built up of three subunits. Two of them are ribophorins - ER transmembrane proteins. Their cytosolic parts bind to the larger subunit of the ribosome (they act as an anchor). The third subunit has the catalytic activity.

Modifications of N-linked oligosaccharides

ER : After an oligosaccharide chain have been added to the protein, the three glucose and one mannose residues are removed by three different enzymes in a fixed order. It occurs in the ER and is a signal that the protein can be transported to the GA for further processing. When the glycosylated protein is improperly folded the glucose residue is again added to it. The oligosaccharide with glucose is recognized by the membrane chaperon protein - calnexin. Glycoprotein bound to calnexin cannot be exported to the Golgi (thus the quality of protein is checked). When the conformation of the protein becomes proper the glucose residue is removed and it is released from the calnexin.
After the processing in the ER the high-mannose type oligosaccharide is formed.
The high-mannose type glycoproteins are transported (in membranous vesicles from the RER to the cis-cisternae of the Golgi.
GOLGI : Processing of glycoproteins in GA depends on their final destination.

There are two main processing pathways :

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