Formation of Neu5Ac

Epimerization to UDP-GalNAc occurs via a specific 4-epimerase (Jacobson and Davidson 1963). This enzyme reaction is probably instrumental in the conversion of exogeneous GalNAc into GlcNAc and subsequently sialic acid, as has been found in several studies (Maley et al. 1968, MacNicollcI al. 1978). 

1 UDP-N-acetylglucosamine 2-epimerase 5.1.3.14 UDP-GlcNAc CMP-Neu5Ac + cytosol ( ) Spivak and Roseman 1966 

2 N-acetylmannosamine kinase 2.7.1.60 ManNAc, ATP 7 ( + ) cytosol Warren and Felsenfeld 1962 

3 N-acetylneuraminate-9-phosphate synthase 4.1.3.20 ManNAc-6-P, PEP 7 — cytosol Watson etal. 1966 

4 N-acetylneuraminate-9-phosphate phosphatase 3.1.3.29 Neu5Ac9P 7 — cytosol Warren and Felsenfeld 1962 

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In conclusion, neuraminic acid 2-pyridyl sulfone, 2-phenyl sulfone or 2-chloro derivatives react with ketones or aldehydes in THF in the presence or absence of HMPA resulting in the near instantaneous and stereospecific formation of Neu5Ac a-C-glycosides. 

On the other hand, the use of 1-cyanoethyl ester derivative 43 for the glycosylation of 48 resulted in /3-exclusive formation of Neu5Ac-(2-6)-Glc sequence 49 in high yield, while in MeCN the production of a-glycoside was preferred [37]. 

In an interesting extension of this work, the Neu5Ac aldolase from E. coli was subjected to directed evolution to expand its catalytic activity for enantiomeric forms of the usual substrates to include A -acetyl-L-mannosamine and L-arabinose with formation of the synthetically important products L-sialic add and L-3-deoxy-L-manno-oct-2-ulosonic add (l-KDO) (163). The evolved Neu5Ac aldolases were characterized by sequence analysis, kinetics, stereoselectivity, and in one case even by an X-ray structure analysis. Again, remote mutations were identified. It is significant... 

On the other hand, the reactions in dichloromethane (see Table 1 entries 9,13, and 14) were of poor stereoselectivity, although the overall yield was high. Furthermore, the formation of (1-glycoside increased with rise of reaction temperature (see Table 1, entry 14). The use of a KDN glycosyl donor 13 that is prepared by treatment [38] of 10 with thiophenol and BF3 etherate in dichloromethane, has also given results similar to those described for Neu5Ac [39]. 

Therefore, it seems most plausible that the stereospecific generation of the p-aceto-nitrilium ion (d) of Neu5Ac from the oxocarbenium ion (c) holds the key of the re-preponderant formation of sialyl glycosides in acetonitrile. Thus, the reactive nucleophiles, such as OH-6 of 17 have a chance to attack on other intermediates (a) (c) before the complete... 

Enzyme catalysis proceeds in four steps. The first step is the binding of the sialoside to the neuraminidase, which causes a considerable distortion of the Neu5Ac pyranose ring from a chair conformation to pseudoboat conformation. The second step leads to the formation of the endocyclic sialosyl cation transition-state via proton transfer from the solvent to the substrate. The final two steps are the formation and release of Neu5Ac [74c]. 

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