NahK/GlmU fusion enzyme: characterization and one-step enzymatic synthesis of UDP-N-acetylglucosamine.

Yafei Zhai, Min Liang, Junqiang Fang, Xiaoyue Wang, Wanyi Guan, Xian-Wei Liu, Peng Wang, Fengshan Wang

National Glycoengineering Research Center, Shandong University, B604 Luneng Keji Dasha, 29-1 Shanda Nanlu, Jinan, 250100 Shandong, People's Republic of China.

Biotechnology letters 2012 Jul

filter terms:

The availability of uridine 5'-diphosphate N-acetylglucosamine (UDP-GlcNAc) is a prerequisite for the GlcNAc-transferase-catalyzed glycosylation reaction. UDP-GlcNAc has already been synthesized using an N-acetylhexosamine 1-kinase (NahK) and a GlcNAc-1-P uridyltransferase (truncated GlmU) and here, a fusion enzyme was constructed with truncated GlmU and NahK. After determination of the optimum catalytic condition (pH 8.0 at 40 °C), the fusion enzyme was used to synthesize UDP-GlcNAc in a single step with a yield of 88 % from GlcNAc, ATP and UTP. Furthermore, a simplified purification method was demonstrated using separation by gel filtration after by-product digestion with alkaline phosphatase. An overall yield of 77 % and a purity of over 90 % were achieved.

Citation

Yafei Zhai, Min Liang, Junqiang Fang, Xiaoyue Wang, Wanyi Guan, Xian-Wei Liu, Peng Wang, Fengshan Wang. NahK/GlmU fusion enzyme: characterization and one-step enzymatic synthesis of UDP-N-acetylglucosamine. Biotechnology letters. 2012 Jul;34(7):1321-6