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Nat Chem Biol. 2015 Jul;11(7):465-71. doi: 10.1038/nchembio.1816. Epub 2015 May 18.

An enzyme-coupled biosensor enables (S)-reticuline production in yeast from glucose.

Author information

1
Department of Bioengineering, University of California, Berkeley, Berkeley, California, USA.
2
1] Department of Biology, Concordia University, Montréal, Québec, Canada. [2] Centre for Structural and Functional Genomics, Concordia University, Montréal, Québec, Canada.

Abstract

Benzylisoquinoline alkaloids (BIAs) are a diverse family of plant-specialized metabolites that include the pharmaceuticals codeine and morphine and their derivatives. Microbial synthesis of BIAs holds promise as an alternative to traditional crop-based manufacturing. Here we demonstrate the production of the key BIA intermediate (S)-reticuline from glucose in Saccharomyces cerevisiae. To aid in this effort, we developed an enzyme-coupled biosensor for the upstream intermediate L-3,4-dihydroxyphenylalanine (L-DOPA). Using this sensor, we identified an active tyrosine hydroxylase and improved its L-DOPA yields by 2.8-fold via PCR mutagenesis. Coexpression of DOPA decarboxylase enabled what is to our knowledge the first demonstration of dopamine production from glucose in yeast, with a 7.4-fold improvement in titer obtained for our best mutant enzyme. We extended this pathway to fully reconstitute the seven-enzyme pathway from L-tyrosine to (S)-reticuline. Future work to improve titers and connect these steps with downstream pathway branches, already demonstrated in S. cerevisiae, will enable low-cost production of many high-value BIAs.

PMID:
25984720
DOI:
10.1038/nchembio.1816
[Indexed for MEDLINE]

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