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Plasmid-Free System and Modular Design for Efficient 5-Aminolevulinic Acid Production by Engineered Escherichia coli

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Abstract

5-Aminolevulinic acid (ALA) is an essential intermediate for many organisms and has been considered for the applications of medical especially in photodynamic therapy of cancer recently. However, ALA production via chemical approach is complicated; hence, microbial manufacturing has received more attentions. In this study, a modular design to simultaneously express ALA synthase from Rhodobacter sphaeroides (RshemA), a non-specific ALA exporter (RhtA), and chaperones was first developed and discussed. The ALA production was significantly increased by coexpressing RhtA and RshemA. Besides, ALA was enhanced by the cofactor pyridoxal phosphate (PLP) which was supplied by expressing genes of pdxK and pdxY or direct addition. However, inclusion bodies of RshemA served as an obstacle; thus, chaperones DnaK and GroELS were introduced to reform the conformation of proteins and successfully improved ALA production. Finally, a plasmid-free strain RrGI, as the robust chassis, was established and a 6.23-fold enhancement on ALA biosynthesis and led to 7.47 g/L titer and 0.588 g/L/h productivity under the optimal cultural condition.

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The data and materials that support the finding of this study are available from the corresponding author upon reasonable request.

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Funding

The study was financially supported by the Ministry of Science and Technology (MOST 108-2221-E-006-004-MY3 and MOST 108-2621-M-006-015) in Taiwan.

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  1. I-Tai Shih and Ying-Chen Yi have equal contribution.

Authors and Affiliations

  1. Department of Chemical Engineering, National Cheng Kung University, Tainan, 70101, Taiwan

    I-Tai Shih, Ying-Chen Yi & I-Son Ng

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  1. I-Tai Shih
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Contributions

ITS and ISN conceived the study. ITS performed all the experiments and original draft investigation. YCY performed part of the analytical method. ISN did methodology validation, supervised the experiments, prepared, review, and edited the manuscript.

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Correspondence to I-Son Ng.

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Shih, IT., Yi, YC. & Ng, IS. Plasmid-Free System and Modular Design for Efficient 5-Aminolevulinic Acid Production by Engineered Escherichia coli. Appl Biochem Biotechnol 193, 2858–2871 (2021). https://doi.org/10.1007/s12010-021-03571-3

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