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

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Abstract

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.

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Acknowledgments

This work was supported by National Natural Science Foundation of China (No. 31000369), China Postdoctoral Science Foundation (No. 20090461238) and, Independent Innovation Foundation of Shandong University (IIFSDU, No. 2009GN038).

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Author notes

  1. Wanyi Guan

    Present address: College of Life Science, Hebei Normal University, Shijiazhuang, 050016, Hebei, People’s Republic of China

Authors and Affiliations

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

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

  2. The State Key Laboratory of Microbial Technology and School of Life Sciences, Shandong University, Jinan, 250100, Shandong, People’s Republic of China

    Yafei Zhai, Min Liang, Xiaoyue Wang, Wanyi Guan & Peng Wang

  3. School of Pharmaceutical Sciences, Shandong University, Jinan, 250100, Shandong, People’s Republic of China

    Junqiang Fang, Xian-wei Liu & Fengshan Wang

Authors
  1. Yafei Zhai
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  2. Min Liang
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  3. Junqiang Fang
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  4. Xiaoyue Wang
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  7. Peng Wang
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Correspondence to Xian-wei Liu.

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Zhai, Y., Liang, M., Fang, J. et al. NahK/GlmU fusion enzyme: characterization and one-step enzymatic synthesis of UDP-N-acetylglucosamine. Biotechnol Lett 34, 1321–1326 (2012). https://doi.org/10.1007/s10529-012-0910-y

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  • DOI: https://doi.org/10.1007/s10529-012-0910-y

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