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Characterisation of a 3-hydroxypropionic acid-inducible system from Pseudomonas putida for orthogonal gene expression control in Escherichia coli and Cupriavidus necator

Hanko, Erik K.R.; Minton, Nigel P.; Malys, Naglis

Authors

Erik K.R. Hanko

Nigel P. Minton



Abstract

3-hydroxypropionic acid (3-HP) is an important platform chemical used as a precursor for production of added-value compounds such as acrylic acid. Metabolically engineered yeast, Escherichia coli, cyanobacteria and other microorganisms have been developed for the biosynthesis of 3-HP. Attempts to overproduce this compound in recombinant Pseudomonas denitrificans revealed that 3-HP is consumed by this microorganism using the catabolic enzymes encoded by genes hpdH, hbdH and mmsA. 3-HP-inducible systems controlling the expression of these genes have been predicted in proteobacteria and actinobacteria. In this study, we identify and characterise 3-HP-inducible promoters and their corresponding LysR-type transcriptional regulators from Pseudomonas putida KT2440. A newly-developed modular reporter system proved possible to demonstrate that PpMmsR/PmmsA and PpHpdR/PhpdH are orthogonal and highly inducible by 3-HP in E. coli (12.3- and 23.3-fold, respectively) and Cupriavidus necator (51.5- and 516.6-fold, respectively). Bioinformatics and mutagenesis analyses revealed a conserved 40-nucleotide sequence in the hpdH promoter, which plays a key role in HpdR-mediated transcription activation. We investigate the kinetics and dynamics of the PpHpdR/PhpdH switchable system in response to 3-HP and show that it is also induced by both enantiomers of 3-hydroxybutyrate. These findings pave the way for use of the 3-HP-inducible system in synthetic biology and biotechnology applications.

Citation

Hanko, E. K., Minton, N. P., & Malys, N. (in press). Characterisation of a 3-hydroxypropionic acid-inducible system from Pseudomonas putida for orthogonal gene expression control in Escherichia coli and Cupriavidus necator. Scientific Reports, 7, https://doi.org/10.1038/s41598-017-01850-w

Journal Article Type Article
Acceptance Date Apr 13, 2017
Online Publication Date May 11, 2017
Deposit Date May 18, 2017
Publicly Available Date May 18, 2017
Journal Scientific Reports
Print ISSN 2045-2322
Electronic ISSN 2045-2322
Publisher Nature Publishing Group
Peer Reviewed Peer Reviewed
Volume 7
Article Number 1724
DOI https://doi.org/10.1038/s41598-017-01850-w
Public URL http://eprints.nottingham.ac.uk/id/eprint/42195
Publisher URL https://www.nature.com/articles/s41598-017-01850-w
Copyright Statement Copyright information regarding this work can be found at the following address: http://creativecommons.org/licenses/by/4.0

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Copyright Statement
Copyright information regarding this work can be found at the following address: http://creativecommons.org/licenses/by/4.0





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