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The rapid high-throughput screening of ω-transaminases via a colorimetric method using aliphatic α-diketones as amino acceptors

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Abstract

ω-Transaminases (ω-TAs) are widely available for the production of chiral amines and unnatural amino acids. Herein, a rapid spectrophotometric method was developed for screening ω-TAs based on the colored products that can be generated from transamination reactions between aliphatic α-diketones and amino donors catalyzed by ω-TAs. The possible mechanism of the formation of the colored product was investigated according to LC-Q-TOF–MS analysis. Among seven diketones, 2,3-butanedione was selected as the most suitable amino acceptor for colorimetric screening of ω-TAs with high efficiency, high sensitivity, and low background interference. Meanwhile, the absorbance of the colored product generated by 2,3-butanedione catalyzed by ω-TAs in this method was linearly correlated with the results by HPLC analysis. This method was also confirmed to effectively screen ω-TA mutants with high activity towards isopropylamine.

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Data availability

All proteins in the present study are available from GenBank via the accession codes. The original data are provided with the paper.

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Acknowledgements

The authors would like to thank the Jiangsu Synergetic Innovation Center for Advanced Bio-manufacture and PAPD to fund this work. For Feng Dan and Shuai Shi, we also thank their work on language checking. The authors also gratefully acknowledge the editors and the reviewers for their useful feedback which improved this paper.

Funding

This work was financially funded by the Jiangsu Synergetic Innovation Center for Advanced Bio-manufacture and PAPD.

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Authors and Affiliations

  1. College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech University, Nanjing, 211816, China

    Kexin Tang, Jiacheng Dong, Zhengheng Zheng, Ting Zhang, Huayi Pan, Honghua Jia, Yan Li & Ping Wei

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  1. Kexin Tang
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  2. Jiacheng Dong
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  3. Zhengheng Zheng
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  8. Ping Wei
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Contributions

K.X. Tang conducted and designed most parts of the experiments. J.C. Dong assisted the first author with HPLC analysis. Z.H. Zheng, T. Zhang, and H.Y. Pan performed activity and data processing. Y. Li. and H.H. Jia. designed and supervised the project. P. Wei was the coordinator of the project. All authors discussed the design and results, commented on the manuscript, and approved the manuscript.

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Correspondence to Honghua Jia or Yan Li.

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Tang, K., Dong, J., Zheng, Z. et al. The rapid high-throughput screening of ω-transaminases via a colorimetric method using aliphatic α-diketones as amino acceptors. Anal Bioanal Chem 415, 1733–1740 (2023). https://doi.org/10.1007/s00216-023-04573-2

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