Enzymatic Synthesis of Sulfated Disaccharides using β-D-Galactosidase-catalyzed Transglycosylation

Abstract

We have established a unique enzymatic approach for obtaining sulfated disaccharides using Bacillus circulans beta-D-galactosidase-catalyzed 6-sulfo galactosylation. When 4-methyl umbelliferyl 6-sulfo beta-D-galactopyranoside (S6Gal beta-4MU) was used as a donor, the enzyme induced transfer of 6-sulfo galactosyl residue to GlcNAc acceptor. As a result, the desired compound 6'-sulfo N-acetyllactosamine (S6Gal beta1-4GlcNAc) and its positional isomer 6'-sulfo N-acetylisolactosamine (S6Gal beta1-6GlcNAc) were observed by HPAEC-PAD, in 49% total yield based on the donor added, and in a molar ratio of 1:3.5. With a glucose acceptor, the regioselectivity was substantially changed and S6Gal beta1-2Glc was mainly produced along with beta-(1-1)alpha, beta-(1-3), beta-(1-6) isomers in 74% total yield. When methyl alpha-D-glucopyranoside (Glc alpha-OMe) was an acceptor, the enzyme also formed mainly S6Gal beta1-2Glc alpha-OMe with its beta-(1-6)-linked isomer in 41% total yield based on the donor added. In both cases, it led to the predominant formation of beta-(1-2)-linked disaccharides. In contrast, with the corresponding methyl beta-D-glucopyranoside (Glc beta-OMe) acceptor, S6Gal beta1-3Glc beta-OMe and S6Gal beta1-6Glc beta-OMe were formed in a low total yield of 12%. These results indicate that the regioselectivity and efficiency on the beta-D-galactosidase-mediated transfer reaction significantly depend on the anomeric configuration in the glucosyl acceptors.

Glycosyl Hydrolases and Glycosyltransferases in the Synthesis of Oligosaccharides

“…The α-anomer of methyl glucoside gave rise to β-1-2 and β-1-6 isomers in 41% yield, while with the β-anomer the yield dropped down to 12% of β-1-3 and β-1-6 disaccharides. These results clearly show the effect of anomeric configuration on the efficiency and selectivity of β-galactosidase transfer reaction also with sulphated sugars [87]. β-Galactosylation of simple alcohols is also an interesting issue.…”

Causes of the Production of Multiple Forms of β-Galactosidase byBacillus circulans

“…4,5,12,13) In particular, -galactosidase-2 does not require high lactose concentrations to produce high yields of oligosaccharides, in contrast to most -galactosidases reported to date. 4) -Galactosidase-2 produces GOS at a yield of about 41% from a 4.56% solution of lactose at 40 C. Hence, during the past 20 years, the commercial enzyme Biolacta Ò has frequently been utilized to synthesize various functional galactosylated compounds [32][33][34][35][36][37][38][39][40][41][42] as well as GOS. 5,9,12,13,43,44) In a later study, Vetere et al 45) isolated a third enzyme, with a molecular mass of 86 kDa, from Biolacta Ò , but they did not study oligosaccharide production.…”

Purification and Characterization of a Novel β-Galactosidase with Transglycosylation Activity from Bacillus megaterium 2-37-4-1

“…Katsumi and Mariko synthesized NeuAcα-(2→3) Galβ-(1→3)-GalNAc, which is regarded as an important tumor marker as a glycopeptides unit, and is called the sialyl T-antigen [4]. Takeomi and Masaki synthesized 6′-sulfoNacetyllactosamine and its positional isomer 6′-sulfoN-acetylisoactosamine, which have been known to play various roles in biological events such as selectin binding, laminin binding, neural cell migration, bacteria binding, and activation of macrophages [5]. Higashiyama and Watanabe synthesized cyclo-{→6)-α-D-Glcp-(1→3)-α-D-Glcp-(1→6)-α-D-Glcp-(1→3)-α-D-Glcp-(1→}, which is a low-calorie sweetener [6].…”