Enzymatic synthesis of novel oligosaccharides from N-acetylsucrosamine using β-fructofuranosidase from Aspergillus oryzae

• Novel oligosaccharides were produced from N-acetylsucrosamine.
• N-Acetyl-1-kestosamine and N-acetylnystosamine were produced.
• Aspergillus oryzae β-fructofuranosidase was used for the oligosaccharide production.
• These oligosaccharides were produced also by using dried A. oryzae mycelia.
• We prepared A. oryzae mycelia suitable for the production of these oligosaccharides.

Mycelia of Aspergillus oryzae NBRC100959 contain 2 types of β-fructofuranosidases, transfructosylation-catalyzing enzyme (βFFaseI), and hydrolysis-catalyzing enzyme (βFFaseII). Using βFFaseI extracted from the mycelia of strain NBRC100959, two novel oligosaccharides consisting of GlcNAc and fructose, β-d-fructofuranosyl-(2→1)-β-d-fructofuranosyl-(2↔1)-2-acetamido-2-deoxy-α-d-glucopyranoside (N-acetyl-1-kestosamine, 1-KesNAc) and β-d-fructofuranosyl-(2→1)-β-d-fructofuranosyl-(2→1)-β-d-fructofuranosyl-(2↔1)-2-acetamido-2-deoxy-α-d-glucopyranoside (N-acetylnystosamine, NysNAc), were synthesized from β-d-fructofuranosyl-(2↔1)-2-acetamido-2-deoxy-α-d-glucopyranoside (N-acetylsucrosamine, SucNAc). We next planned to synthesize 1-KesNAc and NysNAc using A. oryzae mycelia. However, it was thought that the presence of βFFaseII is disadvantageous for the production of these oligosaccharides by βFFaseI, because βFFaseII hydrolyzed 1-KesNAc and NysNAc. We succeeded to produce A. oryzae mycelia containing βFFaseI as the major β-fructofuranosidase, by increasing sucrose concentration in the culture medium. Then, using a dried sample of these A. oryzae mycelia, reaction for the oligosaccharide production was performed. As the results, 190 mg of 1-KesNAc and 60 mg of NysNAc were obtained from 0.6 g of SucNAc. This whole-cell catalysis method facilitates the synthesis of 1-KesNAc and NysNAc because extraction and purification of βFFaseI from mycelia are unnecessary.

Figure optionsDownload as PowerPoint slide