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Journal of Industrial Microbiology & Biotechnology

, Volume 46, Issue 12, pp 1631–1641 | Cite as

Integrating enzyme evolution and high-throughput screening for efficient biosynthesis of l-DOPA

  • Weizhu Zeng
  • Bingbing Xu
  • Guocheng Du
  • Jian Chen
  • Jingwen ZhouEmail author
Biocatalysis - Original Paper
  • 224 Downloads

Abstract

l-DOPA is a key pharmaceutical agent for treating Parkinson’s, and market demand has exploded due to the aging population. There are several challenges associated with the chemical synthesis of l-DOPA, including complicated operation, harsh conditions, and serious pollution. A biocatalysis route for l-DOPA production is promising, especially via a route catalyzed by tyrosine phenol lyase (TPL). In this study, using TPL derived from Erwinia herbicola (Eh-TPL), a mutant Eh-TPL was obtained by integrating enzyme evolution and high-throughput screening methods. l-DOPA production using recombinant Escherichia coli BL21 (DE3) cells harbouring mutant Eh-TPL was enhanced by 36.5% in shake flasks, and the temperature range and alkali resistance of the Eh-TPL mutant were promoted. Sequence analysis revealed two mutated amino acids in the mutant (S20C and N161S), which reduced the length of a hydrogen bond and generated new hydrogen bonds. Using a fed-batch mode for whole-cell catalysis in a 5 L bioreactor, the titre of l-DOPA reached 69.1 g L−1 with high productivity of 11.52 g L−1 h−1, demonstrating the great potential of Eh-TPL variants for industrial production of l-DOPA.

Keywords

Error-prone PCR High-throughput screening Tyrosine phenol lyase Catalytic activity Fed-batch mode 

Notes

Acknowledgements

This work was supported by the National Key Research and Development Program of China (2017YFC1600403), the National Natural Science Foundation of China (31830068, 21822806), the Fundamental Research Funds for the Central Universities (JUSRP51701A), the National First-class Discipline Program of Light Industry Technology and Engineering (LITE2018-08), and the Distinguished Professor Project of Jiangsu Province.

Supplementary material

10295_2019_2237_MOESM1_ESM.docx (285 kb)
Supplementary material 1 (DOCX 285 kb)

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Copyright information

© Society for Industrial Microbiology and Biotechnology 2019

Authors and Affiliations

  1. 1.Key Laboratory of Industrial Biotechnology, Ministry of Education and School of BiotechnologyJiangnan UniversityWuxiChina
  2. 2.National Engineering Laboratory for Cereal Fermentation TechnologyJiangnan UniversityWuxiChina
  3. 3.The Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, School of BiotechnologyJiangnan UniversityWuxiChina
  4. 4.Jiangsu Provisional Research Center for Bioactive Product Processing TechnologyJiangnan UniversityWuxiChina

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