Suddham Singh

Glycosidase-catalysed oligosaccharide synthesis of di-, tri- and tetra-saccharides using the N-acetylhexosaminidase from Aspergillus oryzae and the β-galactosidase from Bacillus circulans

The N-acetylhexosaminidase from Aspergillus oryzae catalyse transfer of N-acetylglucosaminyl and N-acetylgalactosaminyl residues selectively on to the 6-OH group of N-acetylgalactosamine to give the corresponding (1→6)-linked disaccharides 3 and 6 in 26 and 38% yield, respectively. The disaccharide β-D-GlcpNAc-(1→6)-β-D-GalpNAc 3 thus synthesized acts as acceptor for transfer of a β-D-galactosyl residue from the corresponding p-nitrophenyl glycoside on to the 4-OH group of the non-reducing unit to give the ovarian cyst fluid mucin fragment β-D-Galp-(1→4)-β-GlcpNAc-(1→6)-D-GalpNAc 8 in 48% yield together with the tetrasaccharide β-D-Galp-(1→4)-β-D-Gal-(1→4)-β-D-GlcpNAc-(1→6)-D-GalpNAc 9 in 7% yield. With lactose as acceptor, the trisaccharide β-D-Galp-(1→4)-β-D-Galp-(1→4)-D-Glcp 12, a growth factor for human intestinal bifidobacteria, is produced in 52% yield.p

Glycosidase-catalysed synthesis of oligosaccharides: a two-step synthesis of the core trisaccharide of N-linked glycoproteins using the β-N-acetylhexosaminidase and the β-mannosidase from Aspergillus oryzae

Using a partially purified β-mannosidase from Aspergillus oryzae, a β-mannosyl unit is trasferred from p-nitrophenyl β-D-mannopyranoside 5 to di-N-acetylchitobiose 4 to give the core trisaccharide β-D-Manp-(1→4)-β-D-GlcpNAc-(1→4)-D-GlcpNAc 6 of the N-linked glycoproteins.

Kinetic control of regioselectivity in glycosidase-catalysed disaccharide synthesis: preparation of 2-acetamido-4-O-(2-acetamido-2-deoxy-β-D-glucopyranosyl)-2-deoxy-D-glucopyranose (N,n′-diacetylchitobiose) and 2-acetamido-6-O-(2-acetamido-2-deoxy-β-D-glucopyranosyl)-2-deoxy-D-glucopyranose

During transfer of the N-acetyl-β-D-glucosaminyl (2-acetamido-2-deoxy-β-D-glucopyranosyl) residue from p-nitrophenyl N-acetyl-β-D-glucosaminide (p-nitrophenyl N-2-deoxy-β-D-glucopyranoside) on to N-acetyl-D-glucosamine (2-acetamido-2-deoxy-D-glucopyranose) catalysed by the N-acetylhexosaminidase from Aspergillus oryzae, the major isomer formed was found to depend on the time course of the reaction, 1 → 4 transfer predominating while the p-nitrophenyl glycoside was available as donor, but 1 → 6 transfer when the initially-formed 1 → 4 product took over as donor, results that could be interpreted in terms of a constant regioselectivity modulated by selective hydrolysis of the products.

Biotransformations in carbohydrates synthesis. N-Acetylgalactosaminyl transfer on to methyl N-acetyl-β-D-glucosaminide (methyl 2-acetamido-2-deoxy-β-D-glucopyranoside) and methyl N-acetyl-α-D-glucosaminide (methyl 2-acetamido-2-deoxy-α-D-glucopyranoside) catalysed by a β-N-acetylgalactosaminidase from Aspergillus oryzae

Using a crude N-acetylgalactosaminidas from Aspergillus oryzae, the β-N-acetylgalactosaminyl moiety of p-nitrophenyl N-acetylgalactosaminide was transferred to the C-4 and C-6 hydroxy groups of methyl N-acetyl-β-D-glucosaminide and methyl N-acetyl-α-D-glucosaminide and, for the latter, with high efficiency and selectivity for transfer to the C-4 position.

Purification of the β-N-acetylhexosaminidase from Aspergillus oryzae and the β-mannosidases from Helix pomatia and A. oryzae and their application to the enzymic synthesis of the core trisaccharide of the N-linked glycoproteins

The β-N-acetylhexosaminidase from Aspergillus oryzae and the β-mannosidases from Helix pomatia and A. oryzae were purified and used to synthesise the core trisaccharide of N-linked glycoproteins, Manp-(1→4)-β-D-GlcpNAc-(1→4)-D-GlcpNAc.

Glycosidase-catalysed synthesis of α-galactosyl epitopes important in xenotransplantation and toxin binding using the α-galactosidase from Penicillium multicolor

The α-galactosidase from Penicillium multicolor catalyses highly regioselective galactosyl transfer on to mono- and di-saccharide acceptors that have a non-reducing terminal galactose unit to give products containing the α-D-Galp-(1→3)-D-Galp epitope found on pig tissue and which is responsible for the hyperacute rejection response in xenotransplantation of pig organs into man.

Biotransformations in carbohydrate synthesis. N-Acetylgalactosaminyl and N-acetylglucosaminyl transfer onto methyl α- and β-glucosides catalysed by the β-N-acetylhexosaminidase from Aspergillus oryzae

Regioselectivity of N-acetylhexosaminyl transfer onto methyl α- and β-D-glucosides catalysed by the N-acetyl hexosaminidase from Aspergillus oryzae is strongly dependent on the configuration at the anomeric centre of the acceptor: with the β-glucoside, 1,3- and 1,4-transfer products are formed, whereas with the α-glucoside, 1,4-and 1,6-transfer products are obtained.