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Exploring the allosteric mechanism of dihydrodipicolinate synthase by reverse engineering of the allosteric inhibitor binding sites and its application for lysine production

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Abstract

Dihydrodipicolinate synthase (DHDPS, EC 4.2.1.52) catalyzes the first committed reaction of l-lysine biosynthesis in bacteria and plants and is allosterically regulated by l-lysine. In previous studies, DHDPSs from different species were proved to have different sensitivity to l-lysine inhibition. In this study, we investigated the key determinants of feedback regulation between two industrially important DHDPSs, the l-lysine-sensitive DHDPS from Escherichia coli and l-lysine-insensitive DHDPS from Corynebacterium glutamicum, by sequence and structure comparisons and site-directed mutation. Feedback inhibition of E. coli DHDPS was successfully alleviated after substitution of the residues around the inhibitor’s binding sites with those of C. glutamicum DHDPS. Interestingly, mutagenesis of the lysine binding sites of C. glutamicum DHDPS according to E. coli DHDPS did not recover the expected feedback inhibition but an activation of DHDPS by l-lysine, probably due to differences in the allosteic signal transduction in the DHDPS of these two organisms. Overexpression of l-lysine-insensitive E. coli DHDPS mutants in E. coli MG1655 resulted in an improvement of l-lysine production yield by 46 %.

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Acknowledgment

The authors FG, PZ, and JS gratefully thank the financial support from the 973 key basic research program with the project 2011CBA00804 and the Chinese Academy of Sciences with the project KSCX2-EW-G-14-1. CZ and AZE were supported by the German Research Foundation (DFG) through the project ZE 542/6-1.

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Authors and Affiliations

  1. Department of Biopharmaceutic Engineering, School of Chemical Engineering and Technology, Tianjin University, 92 Weijin Road, Nankai District, Tianjin, 300072, China

    Feng Geng

  2. Joint-Lab of Systems Biotechnology, IBB/TUHH and TIB/CAS, Tianjin, 300308, China

    Feng Geng, Zhen Chen, Ping Zheng, Jibin Sun & An-Ping Zeng

  3. Institute for Bioprocess and Biosystems Engineering, Hamburg University of Technology (IBB/TUHH), Denickestrasse 15, D-21073, Hamburg, Germany

    Zhen Chen & An-Ping Zeng

  4. Key Laboratory of Systems Microbial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, China

    Ping Zheng & Jibin Sun

  5. Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences (TIB/CAS), Tianjin, 300308, China

    Ping Zheng & Jibin Sun

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  1. Feng Geng
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  2. Zhen Chen
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  3. Ping Zheng
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  4. Jibin Sun
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  5. An-Ping Zeng
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Correspondence to An-Ping Zeng.

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The authors Feng Geng and Zhen Chen contributed equally.

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Geng, F., Chen, Z., Zheng, P. et al. Exploring the allosteric mechanism of dihydrodipicolinate synthase by reverse engineering of the allosteric inhibitor binding sites and its application for lysine production. Appl Microbiol Biotechnol 97, 1963–1971 (2013). https://doi.org/10.1007/s00253-012-4062-8

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