Chemoenzymatic galactosialylation with integrated cofactor regeneration

Abstract

As a precursor for the chemical synthesis of sialylated oligosaccharides, the trisaccharide glycoside Neu5Ac alpha (2-8)Gal beta (1-4)GlcNAc beta (1-O)-pent-4-ene was synthesized starting from GlcNAc beta (1-O)-pent-4-ene, UDP-glucose and N-acetylneuraminic acid in a one pot reaction employing galactosyltransferase and alpha (2-6)sialyltransferase in a complete cofactor regeneration system.

Chemistry, biochemistry and biology of sialic acids

“…Several of the artificial CMP-sialic acids turned out to be suitable donors for asialo-a, -acid glycoprotein as acceptor with Gal(@lH)GlcNAc a-2,6-sialyltransferase from rat liver as a biocatalyst [3 16,402,4171. Of special interest is the recently reported chemical synthesis of CMP-{NeuSAc(a2-8)NeuSAc} [479]; an attempt to prepare this compound biosynthetically with NeuSAc(a2-8)NeuSAc and CMP-sialic acid synthase failed so far [472]. CMP-9amino-NeuSAc and CMP-NeuSAcNHz have also been used as synthons to prepare fluorescent and photoactivatable analogues [419].…”

“…A third example is the one-pot enzymatic synthesis of Neu5Ac(a24)Gal(~l-4)GlcNAc (and analogues) based on a P-galactosidase-catalyzed galactosylation, using lactose as a donor and GlcNAc as an acceptor, and a pig liver a-2,6-sialyltransferase-catalyzed sialylation with in situ regeneration of CMP-NeuSAc [542]. A fourth example is the enzymatic synthesis of NeuSAc(a24)Gal(fi 14)GlcNAc(fi l-O)pent-4-ene, a precursor for the organic chemical synthesis of higher oligosaccharides [472]. The trisaccharide was synthesized starting from GlcNAc(fi l-O)pent-4-ene, UDP-Gal (in situ generated from UDP-Glc catalyzed by UDP-Gal 4-epimerase), and NeuSAc in a one-pot reaction employing fi-1,4-galactosyltransferase and a-2,6-sialyltransferase in a complete cofactor regeneration system.…”

Applications of enzymes in synthetic carbohydrate chemistry

“…and additional interesting studies are incorporated in the mentioned paper [69]. Parallel studies employing the pent-4-enyl N-acetyllactosaminide with CMPNeu5Ac and a, 2-6 sialyltransferase led to the corresponding a, 2-6-linkage isomer which could be obtained in acceptable yield including the cofactor regeneration [65]. It should be mentioned that also the galactosylation with GaIT could be incorporated into this scheme [65].…”

“…Parallel studies employing the pent-4-enyl N-acetyllactosaminide with CMPNeu5Ac and a, 2-6 sialyltransferase led to the corresponding a, 2-6-linkage isomer which could be obtained in acceptable yield including the cofactor regeneration [65]. It should be mentioned that also the galactosylation with GaIT could be incorporated into this scheme [65]. Further, Neu5Ac could be enzymatically prepared from N-acetylmannosamine and phosphoenol pyruvate (PEP) using Neu5Ac-aldolase, and even this approach could be combined [70] into the SiaT-application for the synthesis of terminally sialylated oligosaccharides.…”

“…This afforded a series of trisaccharide derivatives Neu5Acot2-6Galfll-4GlcNAcfl-l-R. Besides the unblocked trisaccharide a [40], the methyl glycoside b [61], the fl-Asn derivative e [62], the pentapeptide derivative d [63], the allyl e [64], and the pent-4-enyl glycoside f [65] could be prepared in convincing yields.…”