Enzymic oligosaccharide synthesis

This chapter discusses recent examples of enzymic oligosaccharide synthesis on polymer supports. Examples include cases where water insoluble acceptor-polymers of improved accessibility are used with soluble enzymes, and cases where water-soluble acceptor-polymers are used with either insoluble or soluble enzymes. The methodology is useful for preparative-scale reactions and for the study of enzymic properties. 

The challenge for developing methods for synthesizing oligosaccharides, notably for complex oligosaccharides, that can be scaled up to an industrial level is obvious, based on the use of substrates and enzymes that are available at reasonable cost and purity. Strategies for oligosaccharides synthesis include ... 

Several enzymatic routes with sucrase-type enzymes have been established for oligosaccharide synthesis, some of which are applied industrially for food and feed ingredients, and important examples are compiled.5 Convenient routes for new oligosaccharides, however, are rarely available. 

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. 

Glycal assembly on a solid support eliminates the repetitive purifications usually associated with oligosaccharide synthesis. As a method, it has a certain generality as it does not require any specific enzymes or complex starting materials. Both natural and nonnatural sugars may be used in the constructions. 

Oligosaccharide syntheses employing enzymatic reactions would in principle greatly benefit from being performed on a polymer support since the support might effectively facilitate isolation of the final product. Presumably, a water-soluble polymeric support will be preferable to any insoluble support since reaction rates could otherwise become too slow. Glycosidases as synthetic enzymes would be the best candidates to study this type of the enzymic approach to oligosaccharide synthesis. 

P. Wang, G.-J. Shen, Y.-F. Wang, Y. Ichikawa, and C.-H. Wong, Enzymes in oligosaccharide synthesis Active-domain overproduction, specificity study, and synthetic use of an otl,2-mannosyltransferase with regeneration of GDP-Man, J. Org. Chem. 58 3985 (1993). 

Y. Ichikawa, G.-I. Shen, and C.-H. Wong, Enzyme-catalyzed synthesis of sialyl oligosaccharide with in situ regeneration of CMP-sialic acid, J. Am. Chem. Soc. 113 4698 (1991). 

In addition to the high specificity of glycosyltransferases, a major reason for the ordered addition in oligosaccharide synthesis is that these enzymes are found in different membrane-bound compartments. Nascent glycoproteins come in contact with particular glycosyltransferases only when they are in the cellular compartment where the glycosyltransferase is located. 

M. Remaud-Simeon, R. M. Willemot, P. Sargabal, G. Potocki de Montalk, and P. Monsan, Glucansucrases Molecular engineering and oligosaccharide synthesis, J. Mol. Catal. B Enzym., 10 (2000) 117-128. 

Mayer C, Jakeman DL, Mah M et al (2001) Directed evolution of new glycosynthases from Agrobacterium (3-glucosidase a general screen to detect enzymes for oligosaccharide synthesis. Chem Biol 8 437 143... 

Application of Biosynthetic Enzymes in HS and Heparin Oligosaccharide Synthesis... 

Remaud-Simeon M, Willemot R, Sarcabal P, Potocki de Montalk G, Monsan P (2000) Glucansucrases molecular engineering and oligosaccharide synthesis. J Mol Catal B Enzym 10 117-128... 

The fructosyltransferase mined from the commercial preparation Pectinex Ultra SP-L is quite stable towards pH, temperature, and the presence of chemicals. The A aculeatus fructosyltransferase showed a high transferase hydrolase raho that provides it with great potential for oligosaccharide synthesis. The enzyme can be easily immobilized on epoxy-activated supports for better performance. Further studies are required to draw mechanishc conclusions on the nature of the kinetics observed with this fructosyltransferase. 

Additionally, it will be shown that the use of enzymes holds great promise both for the enzymatic or chemoenzymatic construction of deoxygenated donor substrates of glycosyltransferases and for enzymatic oligosaccharide synthesis. 

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