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Metabolic engineering of Escherichia coli for improving l-3,4-dihydroxyphenylalanine (l-DOPA) synthesis from glucose

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Journal of Industrial Microbiology & Biotechnology

Abstract

l-3,4-dihydroxyphenylalanine (l-DOPA) is an aromatic compound employed for the treatment of Parkinson's disease. Metabolic engineering was applied to generate Escherichia coli strains for the production of l-DOPA from glucose by modifying the phosphoenolpyruvate:sugar phosphotransferase system (PTS) and aromatic biosynthetic pathways. Carbon flow was directed to the biosynthesis of l-tyrosine (l-Tyr), an l-DOPA precursor, by transforming strains with compatible plasmids carrying genes encoding a feedback-inhibition resistant version of 3-deoxy-d-arabino-heptulosonate-7-phosphate synthase, transketolase, the chorismate mutase domain from chorismate mutase-prephenate dehydratase from E. coli and cyclohexadienyl dehydrogenase from Zymomonas mobilis. The effects on l-Tyr production of PTS inactivation (PTS− gluc+ phenotype), as well as inactivation of the regulatory protein TyrR, were evaluated. PTS inactivation caused a threefold increase in the specific rate of l-Tyr production (q l-Tyr), whereas inactivation of TyrR caused 1.7- and 1.9-fold increases in q l-Tyr in the PTS+ and the PTS− gluc+ strains, respectively. An 8.6-fold increase in l-Tyr yield from glucose was observed in the PTS− gluc+ tyrR − strain. Expression of hpaBC genes encoding the enzyme 4-hydroxyphenylacetate 3-hydroxylase from E. coli W in the strains modified for l-Tyr production caused the synthesis of l-DOPA. One of such strains, having the PTS− gluc+ tyrR − phenotype, displayed the best production parameters in minimal medium, with a specific rate of l-DOPA production of 13.6 mg/g/h, l-DOPA yield from glucose of 51.7 mg/g and a final l-DOPA titer of 320 mg/l. In a batch fermentor culture in rich medium this strain produced 1.51 g/l of l-DOPA in 50 h.

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Acknowledgments

This work was supported by CONACyT grants 83039 and 126793. AJM was supported by a fellowship from CONACyT. We thank Luz María Martínez, Mercedes Enzaldo, Juan Manuel Hurtado, Mario Trejo and Martín Patiño for technical assistance.

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Authors and Affiliations

  1. Departamento de Ingenierıa Celular y Biocatálisis, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Apdo. Postal 510-3, 62210, Cuernavaca, Morelos, Mexico

    Ana Joyce Muñoz, Georgina Hernández-Chávez, Ramon de Anda, Alfredo Martínez, Francisco Bolívar & Guillermo Gosset

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  1. Ana Joyce Muñoz
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  2. Georgina Hernández-Chávez
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  3. Ramon de Anda
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  4. Alfredo Martínez
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  5. Francisco Bolívar
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  6. Guillermo Gosset
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Correspondence to Guillermo Gosset.

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Muñoz, A.J., Hernández-Chávez, G., de Anda, R. et al. Metabolic engineering of Escherichia coli for improving l-3,4-dihydroxyphenylalanine (l-DOPA) synthesis from glucose. J Ind Microbiol Biotechnol 38, 1845–1852 (2011). https://doi.org/10.1007/s10295-011-0973-0

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  • DOI: https://doi.org/10.1007/s10295-011-0973-0

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