A Chemoenzymatic Synthon Strategy for Synthesizing N-Acetyl Analogues of O-Acetylated N. meningitidis W Capsular Polysaccharide Oligosaccharides

O -Acetylated sialic acid has been found in the Neisseria meningitidis serogroup W (NmW) capsular polysaccharide (CPS) and is a required structural component of clinically used NmW CPS-based polysaccharide and polysaccharide-conjugate vaccines. The role of sialic acid O -acetylation in NmW CPS, however, is not clearly understood. This is partially due to the lack of a precise control of the percentage and the location of O -acetylation which is labile and susceptible to migration. We explore chemoenzymatic synthetic strategies for preparing N -acetylated analogues of O -acetylated NmW CPS oligosaccharides which can serve as structurally stable probe mimics. Substrate specificity studies of NmW CPS polymerase (NmSiaD W ) identified 4-azido-4-deoxy- N -acetylmannosamine (ManNAc4N 3 ) and 6-azido-6-deoxy- N -acetylmannosamine (ManNAc6N 3 ) as suitable chemoenzymatic synthons for synthesizing N -acetyl analogues of NmW CPS oligosaccharides containing 7- O -acetyl- N -acetylneuraminic acid (Neu5,7Ac 2 ) and/or 9- O -acetyl- N -acetylneuraminic acid (Neu5,9Ac 2 ). The synthesis was achieved by NmSiaD W -dependent sequential one-pot multienzyme (OPME) strategy with in situ generation of the corresponding sugar nucleotides from simple monosaccharides or derivatives to form N 3 -oligosaccharides which were converted to the desired NAc-oligosaccharides by an efficient one-step chemical transformation.