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Efficient synthesis of Ala-Tyr by L-amino acid ligase coupled with ATP regeneration system

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Abstract

The multi-enzyme coupling reaction system has become a promising biomanufacturing platform for biochemical production. Tyr is an essential amino acid, but the limited solubility restricts its use. Tyrosyl dipeptide has been paid more attention due to its higher solubility. In this study, an efficient enzymatic cascade of Ala-Tyr synthesis was developed by a L-amino acid ligase together with polyphosphate kinase (PPK). Two L-amino acid ligases from Bacillus subtilis and Bacillus pumilus were selected and applied for Ala-Tyr synthesis. The L-amino acid ligase from B. subtilis (Bs) was selected and coupled with the PPK from Sulfurovum lithotrophicum (PPKSL) for regenerating ATP to produce Ala-Tyr in one pot. In the optimization system, 40.1 mM Ala-Tyr was produced within 3 h due to efficient ATP regeneration with hexametaphosphate (PolyP(6)) as the phosphate donor. The molar yield was 0.89 mol/mol based on the substrates added, while the productivity of Ala-Tyr achieved 13.4 mM/h, which were the highest yield and productivity ever reported about Ala-Tyr synthesis with L-amino acid ligase.

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Funding

This work was supported by the Science and Technology Support Plan for Youth Innovation of Colleges and Universities of Shandong Province (2021KJ109) and the Shandong Provincial Natural Science Foundation (ZR2019MC066).

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Authors and Affiliations

  1. School of Life Science and Bioengineering, Jining University, Jining, 273155, China

    Xiangwei Cui, Xinxin Du, Qiang Zhao, Yanying Hu & Caihong Tian

  2. School of Engineering, Jining University, Jining, 273155, China

    Wenlu Song

Authors
  1. Xiangwei Cui
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  2. Xinxin Du
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  3. Qiang Zhao
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  4. Yanying Hu
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  5. Caihong Tian
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Contributions

All authors contributed to the study’s conception and design. Material preparation and data collection were performed by X. W. Cui, X. X. Du, Q. Zhao, and Y. Y. Hu. Data analysis was performed by C. H. Tian. The first draft of the manuscript was written by W. L. Song, and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.

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Correspondence to Wenlu Song.

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Cui, X., Du, X., Zhao, Q. et al. Efficient synthesis of Ala-Tyr by L-amino acid ligase coupled with ATP regeneration system. Appl Biochem Biotechnol 195, 4336–4346 (2023). https://doi.org/10.1007/s12010-023-04365-5

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  • DOI: https://doi.org/10.1007/s12010-023-04365-5

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