Sialosides sialic acid aldolase

Figure 22 Biosynthesis of sialosides. Sialic acid aldolases catalyze the aldol condensation of W-substituted mannosamines with pyruvate to produce sialic acids. CMP-sialic acid synthetases produce CMP-sialic acid from CTP and sialic acid. Sialic acid is transferred to acceptor molecules by sialyltransferases.

Figure 2 One-pot three-enzyme chemoenzymatic synthesis of sialosides containing sialic acid modifications. In this strategy, mannose or ManNAc derivatives are chemically or enzymatically synthesized. These compounds are then used by a recombinant coli K-12 sialic acid aldolase to obtain sialic acids and their derivatives followed by an N. meningitidis CMP-sialic acid synthetase for the formation of CMP-sialic acids. From which, sialic acids can be transferred to lactose, LacNAc, galactose, GalNAc, or their derivatives by a multifunctional P. muitocida sialyltransferase (PmSTl) or a P, damseia a2,6-sialyltransferase (Pd2,6ST) to form a2,3- or a2,6-linked sialosides in one pot without the isolation of intermediates.

A simplified one-pot two-step enzymatic approach was used by James Paulson s group to produce sialoside derivatives (23). Instead of recycling the CMP-sialic acid, the sialyltransferase donors with modifications at C-5 or C-9 position of NeuSAc were enzymatically synthesized from ManNAc or its C-2 or C-6 modified derivatives, pyruvate, and CTP using a sialic acid aldolase and a CMP-Neu5Ac synthetase/sialyltransferase fusion protein. After removing the protein by membrane filtration, the filtrate containing produced CMP-sialic acid... 

The total synthesis of sialosides by using the chemoenzymatic approach is as follows [74]. Sialic acid itself can be synthesized from ManNAc, mannose, or their derivatives by sialic acid aldolase enzyme through aldol condensation reaction. If ManNAc is chemically or enzymatically modified at C2, C4—C6 positions, sialic acid has structural modifications at C5, C7-C9 positions, respectively. The sialic acids are subsequently activated by a CMP-siahc acid synthetase to form a CMP-sialic acid, which is the donor used by sialyltransferases. Because CMP-sialic acid is tmstable, the CMP-Neu5Ac synthetase is valuable for the preparative enzymatic synthesis of sialosides. In the last steps, the CMP-sialic acid is transferred to galactose or GalNAc terminated glycosides by sialyltransferases to form structurally defined sialosides. Examples are that Chen and co-workers have recently developed a one-pot multienzyme system for the efficient synthesis of a-sialosides (Table 2) [12,76,79]. In this system, recombinant E. coli K-12 sialic acid aldolase catalyzed the synthesis of sialic acid precursors for... 

Our group has recently cloned a truncated Pd2,6ST containing 17-497 amino acid residues as N-hexohistine tagged protein and explored its application in the one-pot three-enzyme system for preparative synthesis of functionalized o2,6-sialosides (25). The tolerance of donor substrate modification by the purified Pd2,6ST was tested using the one-pot three-enzyme system, in which CMP-sialic acid derivatives were generated in situ from sialic acid precursors by the aldolase and NmCSS. An extremely relaxed donor substrate specificity was observed for Pd2,6ST. The preparative-sacle reactions were then carried out at... 

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