ER-Bound Steps in the Biosynthesis of G Protein-Coupled Receptors - GPCR Signalling Complexes: Synthesis, Assembly, Trafficking and Specificity - page 6

a

ER lumen

Cytoplasm

b

c

S

S

oligosaccharyltransferase

core glycan

 -helix and
stop transfer sequence

 -helix and signal anchor

signal peptide translocator channel

Fig. 1.2 Co-translational modification of the polypeptide chain of G protein coupled receptors.
( a ) and ( b ) Transfer of a core glycan (glucose 3 mannose 9 -N-acetylglucoside 2 ) by oligosaccharyl-
transferase (OST) to an asparagine residue present in the N-terminus of the receptor. Typically, more
than one asparagine residue on the luminal side is substrate to co-translational N-glycosylation.
Shown is a signal peptide-less receptor ( a ) and a receptor with signal peptide ( b ) to illustrate a
potential difference in the role of the N-glycan during folding of the N-terminus. ( c ) Disul fi de bond
formation in the prototypical position linking a cysteine at the C-terminal end of helix III and extra-
cellular loop 2. Because the bond forms spontaneously and is covalent, dissolving an incorrect bond
requires enzymatic assistance from thioredoxin-containing enzymes ( e.g. , PDI) in the ER lumen

1.1.3 Glycosylation

The functional complex that comprises the active ribosome and the hetero-trimeric
Sec61 channel complex also includes the hetero-octameric oligosaccharyltransferase
(OST). By association with the ribosome OST scans unfolded polypeptide emerging
from the Sec61 channel for the presence of a glycosylation site represented by an
Asn-X-Ser/Thr tripeptide motif (also designated as sequon). OST (precisely: its
SST3 subunit) catalyzes transfer of the preassembled high-mannose core oligosac-
charide [glucose 3 -mannose 9 -N-acetylglucoside 2 ] from the membrane-bound lipid
donor dolichol pyrophosphate to establish an N-glycosidic link to the asparagine
side chain (Fig. 1.2 ). After glycan attachment, the terminal glucose is removed by
glucosidase I (Glc I), followed by removal of the second glucose by Glc II (Pult
et al. 2011 ) . This generates [glucose 1 -mannose 9 -N-acetylglucoside 2 ], a high affinity