HALF-LIFE EXTENSION THROUGH O-GLYCOSYLATION - Fusion Protein Technologies for Biopharmaceuticals: Applications and Challenges - page 196

N-linked O-linked

SA SA

Gal Gal

GlcNAc GlcNAc

Man Man

SA

Man

Gal

GlcNAc

GalNAc

SA

GlcNAc

Ser/Thr

Asn -X-Ser/Thr

FIGURE 13.1 Structure of typical N- and O-linked oligosaccharide chains.

terminating with negatively charged sialic acid residue
(Figure 13.1).
Protein glycosylation is believed to be the most compli-
cated post-translational event. COHs are added to proteins in
two organelles: the ER and the Golgi apparatus. COHs
addition to proteins occurs both co- and post-translationally.
The RNA coding the protein sequence enters the cytoplasm
where it binds to ribosomes, which are the site for protein
synthesis. Ribosomes bind to the ER and the nascent protein
chain enters the lumen and a core oligosaccharide is added to
the protein. Further additions of monosaccharides are pre-
formed in the lumen until a final core mannose structure has
been added. The ER lumen contains high concentrations of
molecular chaperones to assist protein folding. Additional
COH modifications (post-translational) are made as the
protein moves from the lumen of the ER to the Golgi
apparatus. Here, terminal COH modification is completed.
The Golgi does not contain molecular chaperons since
protein folding is complete when the proteins arrive. Rather
they have high concentrations of membrane bound enzymes,
including glycosidases and glycosyltransferases.

connection between O-GlcNAc and the development of insulin
resistance involves the hexosamine biosynthesis pathway
(HBP), HBP being the metabolic pathway leading to synthesis
of UDP-GlcNAc, the direct donor substrate for O-GlcNAc. The
role of O-GlcNAc in striated muscle has to be considered in a
larger extent than its implication in insulin resistance. Indeed,
the activity of O-GlcNAc transferase is twofold to fourfold
higher in skeletal muscle and heart than in liver, therefore
suggesting an important role of this post-translational modifi-
cation in muscle “normal” physiology [1-3]. More generally,
different experiments have showed that blocking or reducing
O-GlcNAc increased the sensitivity of cells to stress, causing a
decrease in cell survival, whereas an increase in O-GlcNAc
level protected cells against stress [4].
Other two main functions of O-GlcNAc were described.
One function involves secretion to form components of the
extracellular matrix, adhering one cell to another by inter-
actions between the large sugar complexes of proteoglycans.
The other main function is to act as a component of mucosal
secretions, and it is the high concentration of COHs that
tends to give mucus its “slimy” feel.
One apparently universal consequence of O-linked oli-
gosaccharide chains is relatively resistance to proteases of
O-glycosylated regions in glycoproteins. The most likely
explanation for protease resistance is simply that the
attached COH blocks access to the peptide core since these
same sequences are quite susceptible to proteases in the
absence of attached COH. The second consequence of
O-glycosylation is the induction of a specific conformation [5].
The human chorionic gonadotropin (hCG) b -subunit is
distinguished from the other human

13.2 THE ROLE OF O-LINKED
OLIGOSACCHARIDE CHAINS IN
GLYCOPROTEIN FUNCTION

Many reports suggest that O-GlcNAc might be implicated in
pathological conditions such as cancer, cardiovascular, or
neurodegenerative disorders. Several reports closely link
O-GlcNAc to glucose toxicity and insulin resistance. This tight

b -subunits of the