Simple and large-scale production of N-acetylneuraminic acid from N-acetyl-d-glucosamine and pyruvate using N-acyl-d-glucosamine 2-epimerase and N-acetylneuraminate lyase

N-Acetylneuraminate lyase and N-acyl-d-glucosamine 2-epimerase had been cloned and overexpressed in Escherichia coli. Simultaneous use of these two enzymes and feeding of appropriate amounts of pyruvate to the reaction mixture made possible the high conversion of N-acetylneuraminic acid (Neu5Ac) from N-acetyl-d-glucosamine (GlcNAc) with a 77% conversion rate on a molar basis. As a result, 29 kg of Neu5Ac was obtained from 27 kg of GlcNAc. The product was recovered by direct crystallization, and verified as identical to authentic Neu5Ac.