Development of a Vanillate Biosensor for the Vanillin Biosynthesis Pathway in .

ACS Synth Biol

Department of Chemical Engineering , Massachusetts Institute of Technology, 77 Massachusetts Avenue , Cambridge , Massachusetts 02139 , United States.

Published: September 2019

The engineered vanillin biosynthesis pathway constructed in is industrially relevant but limited by the reaction catalyzed by catechol -methyltransferase, which is intended to catalyze the conversion of protocatechuate to vanillate. To identify alternative -methyltransferases, we constructed a vanillate sensor based on the VanR-VanO system. Using an promoter library, we achieved greater than 14-fold dynamic range in our best rationally constructed sensor. We found that this construct and an evolved variant demonstrate remarkable substrate selectivity, exhibiting no detectable response to the regioisomer byproduct isovanillate and minimal response to structurally similar pathway intermediates. We then harnessed the evolved biosensor to conduct rapid bioprospecting of natural catechol -methyltransferases and identified three previously uncharacterized but active -methyltransferases. Collectively, these efforts enrich our knowledge of how biosensing can aid metabolic engineering and constitute the foundation for future improvements in vanillin pathway productivity.

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Source
http://dx.doi.org/10.1021/acssynbio.9b00071DOI Listing

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