Molecular Biology of the Cell by Bruce Alberts, Alexander Johnson, Julian Lewis, David Morgan, Martin Raff, Keith Roberts, Peter Walter by by Bruce Alberts, Alexander Johnson, Julian Lewis, David Morg

684 Chapter 12: Intracellular Compartments and Protein Sorting
oligosaccharyl transferase, which has its active site exposed on the lumenal side
of the ER membrane; this explains why cytosolic proteins are not glycosylated
in this way. A special lipid molecule called dolichol anchors the precursor oligosaccharide
in the ER membrane. The precursor oligosaccharide is transferred
to the target asparagine in a single enzymatic step immediately after that amino
acid has reached the ER lumen during protein translocation. The precursor oligosaccharide
is linked to the dolichol lipid by a high-energy pyrophosphate bond,
which provides the activation energy that drives the glycosylation reaction (Figure
12–47B). One copy of oligosaccharyl transferase is associated with each protein
translocator, allowing it to scan and glycosylate the incoming polypeptide chains
efficiently.
The precursor oligosaccharide is built up sugar by sugar on the membrane-bound
dolichol lipid and is then transferred to a protein. The sugars are
first activated in the cytosol by the formation of nucleotide (UDP or GDP)-sugar
intermediates, which then donate their sugar (directly or indirectly) to the lipid in
an orderly sequence. Part way through this process, the lipid-linked oligosaccharide
is flipped, with the help of a transporter, from the cytosolic to the lumenal
side of the ER membrane (Figure 12–48).
All of the diversity of the N-linked oligosaccharide structures on mature glycoproteins
results from the later modification of the original precursor oligosaccharide.
While still in the ER, three glucoses (see Figure 12–47) and one mannose are
quickly removed from the oligosaccharides of most glycoproteins. We shall return
to the importance of glucose trimming shortly. This oligosaccharide “trimming,”
or “processing,” continues in the Golgi apparatus, as we discuss in Chapter 13.
The N-linked oligosaccharides are by far the most common oligosaccharides,
being found on 90% of all glycoproteins. Less frequently, oligosaccharides are
linked to the hydroxyl group on the side chain of a serine, threonine, or hydroxylysine
amino acid. A first sugar of these O-linked oligosaccharides is added in the
ER and the oligosaccharide is then further extended in the Golgi apparatus (see
Figure 13–32).
ER LUMEN
“FLIPPING” IN
MEMBRANE
(Man) 5 (GlcNAc) 2
Man P
4 X
P
(Man) 9 (GlcNAc) 2 P P
3 X
(Glc) 3 (Man) 9 (GlcNAc) 2
P
Glc
P
P
P
P
P
lipid bilayer of
ER membrane
dolichol
dolichol
dolichol
dolichol
dolichol
dolichol
dolichol
dolichol
dolichol
dolichol
dolichol
CYTOSOL
P
P
P
P
P
CTP
CDP
2 UDP GlcNAc
UDP + UMP
(GlcNAc) 2
5 GDP
5 GDP
Man
(GlcNAc) 2 (Man) 5
mannose donor made from
dolichol phosphate
and GDP-mannose
glucose donor made from
dolichol phosphate
and UDP-glucose
Figure 12–48 Synthesis of the lipidlinked
precursor oligosaccharide
in the rough ER membrane. The
oligosaccharide is assembled sugar by
sugar onto the carrier lipid dolichol (a
polyisoprenoid; see Panel 2–5, pp. 98–99).
Dolichol is long and very hydrophobic: its
22 five-carbon units can span the thickness
of a lipid bilayer more than three times, so
that the attached oligosaccharide is firmly
anchored in the membrane. The first sugar
is linked to dolichol by a pyrophosphate
bridge. This high-energy bond activates
the oligosaccharide for its eventual transfer
from the lipid to an asparagine side chain
of a growing polypeptide on the lumenal
side of the rough ER. As indicated, the
synthesis of the oligosaccharide starts on
the cytosolic side of the ER membrane
and continues on the lumenal face after
the (Man) 5 (GlcNAc) 2 lipid intermediate is
flipped across the bilayer by a transporter
(which is not shown). All the subsequent
glycosyl transfer reactions on the lumenal
side of the ER involve transfers from
dolichol-P-glucose and dolichol-Pmannose;
these activated, lipid-linked
monosaccharides are synthesized from
dolichol phosphate and UDP-glucose or
GDP-mannose (as appropriate) on the
cytosolic side of the ER and are then
flipped across the ER membrane. GlcNAc
= N-acetylglucosamine; Man = mannose;
Glc = glucose.