Glycosylations with Transferases

Relatively early reports (1980-1982) from Barker and his group described galactosylation77 and fucosylation78 with soluble transferases. 

Sialyl residues in oligosaccharides are introduced by the reaction of cyti-dine monophosphate-V-acetylneuraminic acid (49) as the sugar donor with the appropriate substrate, in the presence of specific transferases. Three of these have been utilized in syntheses which may be considered to be preparative. None are readily available. The most common, which we have called STA (see Table I), catalyzes the transfer of a 5-acetamido-3,5-di-deoxy-D-g/ycm -a -D-ga/arfo-2-hexulopyranosonic acid unit (the a-D-pyra-nose form of JV-acetylneuraminic acid) to the primary position of D-galactose in a JV-acetyllactosamine residue.86 This enzyme also transfers vV-acetyl-9-O-acetylneuraminic acid (20) and V-glycolylneuraminic acid (12) from the corresponding cytidine monophosphate derivatives.16 The commercial enzyme is rather expensive, but pork liver from a butcher is a 

The second transferase (STB see Table I) is also commercially available, and is still more expensive. It catalyses the transfer of N-acetylneuraminic acid to 0-3 of D-galactose in the terminal residue / -D-Galp-( 1 — 3)-D-Gal-NAc.88 The third one (STC), so far does not appear to be at all easily available. It catalyzes the transfer of a JV-acetylneuraminic acid residue to 0-3 of D-galactose in a / -D-Gal/ -[ 1 — 3(4)]-/J-d-G1c/ NAc residue.18,89 

Most sialylations so far reported have been achieved with soluble transferases, and seldom on a more than 20-/imol scale (see Table IX), with the intention to prepare and describe sequences present in glycoproteins and glycolipids. Trisaccharide -D-Neu5Ac-(2— 3)-/J-D-Galp-(l — 3)-/J-d- 

GlqpNAc-( 1 — OMe) was prepared with two different transferases, STB and STC. In our view, the greater efficiency of STB in this preparation deserves further investigation, as the reverse observation might have been expected in view of the known specificities of these enzymes. 

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High selectivity and substrate specificity of glycosyl transferases make them valuable catalysts for special linkages in polymer-supported synthesis. There is, however, still a rather limited set of enzymes available to date, and the need to synthesize a variety of natural and non-natural oligosaccharides prevails. Particularly with regard to combinatorial approaches, chemical solid-phase oligosaccharide synthesis promises to meet the demands most effectively. 

A hexaglycine spacer was attached to the solid support to give a substitution of 0.2 mmol g of dry silica and the excess amino groups were then capped using acetic anhydride. In the next step a selectively cleavable, a-chymotrypsin sensitive, phenylalanine ester (2) was implemented for the release of the products from the solid support under mild conditions. Subsequently it was transformed to (3) followed by reactions with glycosyl transferases to yield (4). Finally, the desired glycopeptide was cleaved from the solid support in high yield by treatment of (4) with a-chymotrypsin (Scheme 10.1). 

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