Sialylated trisaccharides, synthesis

Y. Ichikawa, J. L.-C. Liu, G.-J. Shen, and C.-H. Wong, A highly efficient multienzyme system for the one-step synthesis of sialyl trisaccharide In situ generation of sialic acid and TV-acetyllactosamine coupled with regeneration of UDP-glucose, UDP-galactose, and CMP-sialic acid, J. Am. Chem. Soc. 113 6300 (1991). 

SCHEME 5.16 Use of an ALacetyl-4-0,5-ALoxazolidinone-protected sialyl phosphate in a one-pot trisaccharide synthesis. 

Synthesis of Sialyated Trisaccharides. We have used a combination of glycosidases and glycosyltranferases to prepare sialylated trisaccharides (Equations 8-9). The enzyme P-galactosidase from bovine... 

The immobilized enzyme was used for repeated synthesis of the above sialylated trisaccharides without adding detergent to the system, which was used in previous synthesis with the soluble enzyme (42), The use of detergent complicates isolation of the products. The stability of the enzyme preparation was good and no decrease of activity was observed on repeated preparative synthesis. Recently, immobilization to CNBr-activated agarose of the a2-6sialyltransferase was reported (50),... 

Based on the abiHty of the E. coli NeuA to produce 9-glycosylated KDN 47 from a disaccharide 46 aldol acceptor, the synthesis of the unusual a-2,3- and a-2,6-linked sialylated trisaccharide 48 and tetrasaccharide 49 was reported, in which sialic acids are hnked in both the terminal and reducing positions (Scheme 17.17) [43]. Each transformation was conducted as a one-pot two-step cascade using a combination of CSS and one of the complementary SiaT types. 

Scheme 13. Enzymatic synthesis of a sialylated trisaccharide derivative, by stepwise sugar-chain elongation, using a water-soluble chymotrypsin-sensitive glycopolymer. Reagents i) 1. HEPES buffer pH 6, UDP-Gal (1.5 eq.), GT, 37 °C, 24 h 2. cacodylate buffer pH 7.4, ST6, CIAP, CMP-NeuAc (1.5 eq.), 37°C, 48 h, gel filtration on Sephadex G-25, q.y. ii) 1. a-chymotrypsin, Tris HCl buffer pH 7.8, 40 °C, 24 h 2. gel filtration on Sephadex G-15, 87%.

Lex-antigenic trisaccharide (a-L-Fuc(l-3)-[p-D-Gal-(l-4)-p-D-GlcNAc]) and its sialylated structure (SLex) are terminal components of a nnmber of glycoconjugates on cell surfaces [31]. SLex serves as a ligand for the endothelial leukocyte molecule-1 (E-selectin) [32], which mediates the initial stages of adhesion of leukocytes to activated endothelial cells, and pays a critical role during inflammatory responses [33]. Lex-based carbohydrates have shown promise in therapeutic investigations related to the inflammatory process. As a result, extensive efforts have been directed toward the synthesis of SLex and related molecules [34 also see Chap. 15],... 

The preparation of trisaccharide 63 illustrates the activation and enzymic coupling of the 9-acetate of vV-acetylneuraminic acid, This involves the utilization of enzymes in a cascade of reactions which probably do not occur in cells (a) synthesis of Neu5,9Ac2 from the 6-acetate of vV-acetylmannosa-mine with the catabolic sialyl aldolase, (b) activation with CMPNeu5Ac synthetase, and (c) coupling. Acetylation in cells seems posterior to coupling. Terminal nonreducing vV-acetyl-9-O-acetylneuraminic acid residues appear... 

Following the enzymatic synthesis of CMP-Neu5Ac, isolated a2-6-sialyl-transferases could be used to introduce sialic acid to the N-acetyl lactosamine derivates a-f in position 6. This afforded a series of trisaccharide derivates Neu5Aca2-6 Gal l-4GlcNAc l-R. The unblocked trisaccharide a [55], the methyl glycoside b [56], the -Asn derivative c [57], the pentapeptide derivative d [58], the allyl e [59], and the pent-4-enyl glycoside f [48] can be prepared in convincing yields (Fig. 6). 

SCHEME 15.16 Synthesis of 2,3-sialyl-T antigen building blocks by P-glycosylation of Tn antigen 144, or by a-glycosylation of serine and threonine building blocks with trisaccharide donor 150. 

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