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Production of glutamine synthetase in Escherichia coli using SUMO fusion partner and application to l-glutamine synthesis

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Abstract

l-glutamine (Gln) is an important conditionally necessary amino acid in human body and potential demand in food or medicine industry is expected. High efficiency of l-Gln production by coupling genetic engineered bacterial glutamine synthetase (GS) with yeast alcoholic fermentation system has been developed. We report here first the application of small ubiquitin-related modifier (SUMO) fusion technology to the expression and purification of recombinant Bacillus subtilis GS. In order to obtain GS with high Gln-forming activity, safety and low cost for food and pharmaceutics industry, 0.1% (w/v) lactose was selected as inducer. The fusion protein was expressed in totally soluble form in E. coli, and expression was verified by SDS–PAGE and western blot analysis. The fusion protein was purified to 90% purity by nickel nitrilo-triacetic acid (Ni–NTA) resin chromatography with a yield of 625 mg per liter fermentation culture. After the SUMO/GS fusion protein was cleaved by the SUMO protease, the cleaved sample was reapplied to a Ni–NTA column. Finally, about 121 mg recombinant GS was obtained from 1 l fermentation culture with no less than 96% purity. The recombinant purified GS showed great transferase activity (23 U/mg), with 25 U recombinant GS in a 50 ml reaction system, a biosynthesis yield of 27.5 g/l l-Gln was detected by high pressure liquid chromatography (HPLC) or thin-layer chromatography. Thus, the application of SUMO technology to the expression and purification of GS potentially could be employed for the industrial production of l-Gln.

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Acknowledgments

This work was supported by the special funding from NJNU for talent faculty and Jiangsu superior discipline. We thank zhili Liu, associate professor, for performing the HPLC analysis.

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Authors and Affiliations

  1. Jiangsu Province Key Laboratory for Molecular and Medicine Biotechnology, College of Life Science, Nanjing Normal University, No. 1 Wenyuan Road, 210046, Nanjing, Jiangsu, People’s Republic of China

    Qi Wang, Cui Min, Tingting Yan, Hefang Pu, Yinqiang Xin, Shuangquan Zhang & Zhimin Yin

  2. State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences, Nanjing University, 210093, Nanjing, People’s Republic of China

    Lan Luo

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  1. Qi Wang
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  2. Cui Min
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  3. Tingting Yan
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  4. Hefang Pu
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  5. Yinqiang Xin
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  6. Shuangquan Zhang
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  7. Lan Luo
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  8. Zhimin Yin
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Correspondence to Lan Luo or Zhimin Yin.

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Wang, Q., Min, C., Yan, T. et al. Production of glutamine synthetase in Escherichia coli using SUMO fusion partner and application to l-glutamine synthesis. World J Microbiol Biotechnol 27, 2603–2610 (2011). https://doi.org/10.1007/s11274-011-0733-3

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  • DOI: https://doi.org/10.1007/s11274-011-0733-3

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