Carbohydrate-targeted optimization of therapeutic gonadotropins - Carbohydrate Chemistry: Chemical and Biological Approaches - page 476

biopharmaceutical industry because most expression systems fail to
synthesize highly sialylated glycans. 162 Thus, such optimization also
applies to gonadotropins with an additional benefit for them in that
sialylated glycoforms are more active at the target receptor.

6.2.1 Sialyltransferases. In eukaryotic cells, addition of terminal
sialic acid to glycoconjugates is carried out by specific enzymes called
sialyltransferases (STs). STs catalyze the transfer of a sialic acid from the
activated nucleotide sugar donor CMP-N-acetylneuraminic acid (CMP-
Neu5Ac) to the terminal nonreducing position of oligosaccharide chains
either of glycoproteins or glycolipids.

6.2.1.1 Classification of sialyltransferases. To date, 20 different STs
acting on glycoproteins and/or glycolipids have been cloned and char-
acterized in humans. They are classified into four groups, depending on
the nature of their sugar acceptor and the type of linkage they form:
ST6Gal, ST6GalNAc, ST3Gal and ST8Sia. 163 The various ST6Gal and
ST6GalNAc synthesize a2,6-linkages, ST3Gal form a2,3-linkages and
ST8Sia catalyze the formation of a2,8-linkage. The ST6Gal group is
composed of two members (ST6Gal-I and ST6Gal-II) that catalyze the
transfer of Neu5Ac residues to terminal galactose (Gal) residue of type 2
disaccharide (Galb1-4GlcNAc), and potentially to the GalNAc residue of
LacdiNAc (GalNAcb1-4GlcNAc). The ST6GalNAc group consists in six
different members (ST6GalNAc-I to -VI) that catalyze the transfer to a
GalNAc residue present in O-glycoproteins (ST6GalNAc-I, -II and -IV) or in
glycolipids (ST6GalNAc-III, -V and -VI). The six members of the ST3Gal
group (ST3Gal-I to -VI) catalyze the transfer of Neu5Ac on terminal
galactose residues found in glycoproteins or glycolipids. The six enzymes
of the ST8Sia group (ST8Sia-I to -VI) mediate the transfer of Neu5Ac
on another terminal Neu5Ac residue also present in glycoproteins or
glycolipids. 164

6.2.1.2 Structure of sialyltransferases. STs evolved secondary to many
gene duplications which occurred early in vertebrate evolution. In add-
ition, the appearance of the four main ST groups has occurred among
ancestors of the present invertebrates. 163 All vertebrate STs have a similar
architecture. They are type II transmembrane proteins predominantly
resident in the trans-Golgi compartment and are constituted of a short
N-terminal cytoplasmic tail, a single transmembrane domain (16-20
residues) followed by a stem region of variable length (20-200 amino
acids) and a large C-terminal catalytic domain facing the Golgi lumen
(Fig. 7). 165
Sequence analysis revealed that the length and amino acid com-
position of catalytic domains are relatively well conserved among species.
Significant homology is present as four conserved peptide sequences
designated as sialylmotifs L (large), S (small), 166,167 III (third portion in

the sequence) 168 and VS (very small) (Fig. 7A). 168,169
The sialylmotif L consists in 48-49 amino acids in the middle of the
luminal catalytic domain with eight invariant residues including one
cysteine residue and exhibits W60-70% identity among homologous