In situ Generation of Sugar Nucleotides

In comparison with the fully chemical synthesis of nucleotide-activated donor sugars the chemo-enzymatic or enzymatic approaches are distinguished by fewer protecting group manipulations and high chemo- and stereoselectivity. This simplifies purification protocols significantly. These features reduce the number of synthetic steps and increase overall yields. 

The fully enzymatic preparation of sugar nucleotides, however, requires the availability of a series of enzymes. In addition, their mutual interactions presupposes exact knowledge of their physiological properties by the user this, in particular, makes chemists hesitate to use biocatalysts. Not all the involved enzymes may tolerate altered, non-natural substrates. A combined chemo-enzymatic approach therefore offers the possibility of combining the advantages of the chemical synthesis and those of the enzymatic preparation, e.g. to prepare non-natural nucleotide activated substrates. 

Especially noteworthy is the use of sucrose, Glca(l-2)Fru, as the ultimate source of UDP-Gal. In a first step sucrose synthase from rice cleaves the disaccharide and forms UDP-Glc in the presence of UDP. UDP-Glc is subsequently epimerized at the 4-position by UDP galactose epimerase to give the required UDP-Gal. This donor substrate is first used by P(l-4)galactosyl transferase to produce the N-acetyllactosamine intermediate and then by a(l-3)galactosyl transferase to gal- 

This task can also be achieved by use of microorganisms—the so called wholecell synthesis—provided the microorganisms do not metabolize the product oligosaccharide. 

The practicability of such cyclic multi-enzyme preparations has been confirmed, e.g. for the synthesis of A -acetyllactosamine or the tetrasaccharide, sialyl Lewis, containing four different monosaccharide units. 

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Enzymatic synthesis using partiai in-situ generation of sugar nucleotides... 

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