Enzymatic transfer of sialic acids modified at C-5 employing four different sialyltransferases.

We present kinetic studies on the enzymatic transfer of several synthetic sialic acid analogues, modified at C-5, to distinct glycoprotein glycans by sialyltransferases differing in acceptor- and linkage-specificity. Biochemical properties of sialic acids were modified by introducing formyl-, trifluoroacetyl-, benzyloxy-carbonyl-, and aminoacetyl-groups to the amino group at C-5 of neuraminic acid. The latter substitution renders the corresponding alpha-glycoside resistant towards sialidases. The respective CMP-sialic acid analogues were prepared by CMP-sialic acid synthase with a yield of 13-55%. The kinetic parameters of several sialyltransferases for the 5-substituted CMP-glycosides differed significantly. Relative to parent CMP-NeuAc, reaction rates of human- and rat liver Gal beta 1, 4GlcNAc alpha 2,6-sialyltransferases ranged from 50 to 170%, of GalNAc alpha 2,6-sialyltransferases from 40-140%, and of Gal beta 1,3Gal-NAc alpha 2,3-sialyltransferase from 20-50%. Resialylation of asialo-alpha 1-acid glycoprotein by 5-N-formyl- and 5-N-aminoacetyl-neuraminic acid employing rat liver Gal beta 1,4GlcNAc alpha 2,6-sialyltransferase proceeded to about 80% of galactose sites which is identical to the extent achieved with parent NeuAc. According to our data, neosialoglycoconjugates which carry sialic acids modified at the N-acetyl group can be prepared for structure-function analysis, as this position seems crucial for recognition of adhesion proteins and influenza viruses.