Characterisation of a 3-hydroxypropionic acid-inducible system from Pseudomonas putida for orthogonal gene expression control in Escherichia coli and Cupriavidus necator
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 ov...
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| Format: | Article |
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Nature Publishing Group
2017
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| Online Access: | https://eprints.nottingham.ac.uk/42195/ |
| _version_ | 1848796442126188544 |
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| author | Hanko, Erik K.R. Minton, Nigel P. Malys, Naglis |
| author_facet | Hanko, Erik K.R. Minton, Nigel P. Malys, Naglis |
| author_sort | Hanko, Erik K.R. |
| building | Nottingham Research Data Repository |
| collection | Online Access |
| description | 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. |
| first_indexed | 2025-11-14T19:48:03Z |
| format | Article |
| id | nottingham-42195 |
| institution | University of Nottingham Malaysia Campus |
| institution_category | Local University |
| last_indexed | 2025-11-14T19:48:03Z |
| publishDate | 2017 |
| publisher | Nature Publishing Group |
| recordtype | eprints |
| repository_type | Digital Repository |
| spelling | nottingham-421952020-05-04T18:45:28Z https://eprints.nottingham.ac.uk/42195/ 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 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. Nature Publishing Group 2017-05-11 Article PeerReviewed Hanko, Erik K.R., Minton, Nigel P. and Malys, Naglis (2017) 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 . 1724/1-1724/13. ISSN 2045-2322 https://www.nature.com/articles/s41598-017-01850-w doi:10.1038/s41598-017-01850-w doi:10.1038/s41598-017-01850-w |
| spellingShingle | Hanko, Erik K.R. Minton, Nigel P. Malys, Naglis Characterisation of a 3-hydroxypropionic acid-inducible system from Pseudomonas putida for orthogonal gene expression control in Escherichia coli and Cupriavidus necator |
| title | Characterisation of a 3-hydroxypropionic acid-inducible system from Pseudomonas putida for orthogonal gene expression control in Escherichia coli and Cupriavidus necator |
| title_full | Characterisation of a 3-hydroxypropionic acid-inducible system from Pseudomonas putida for orthogonal gene expression control in Escherichia coli and Cupriavidus necator |
| title_fullStr | Characterisation of a 3-hydroxypropionic acid-inducible system from Pseudomonas putida for orthogonal gene expression control in Escherichia coli and Cupriavidus necator |
| title_full_unstemmed | Characterisation of a 3-hydroxypropionic acid-inducible system from Pseudomonas putida for orthogonal gene expression control in Escherichia coli and Cupriavidus necator |
| title_short | Characterisation of a 3-hydroxypropionic acid-inducible system from Pseudomonas putida for orthogonal gene expression control in Escherichia coli and Cupriavidus necator |
| title_sort | characterisation of a 3-hydroxypropionic acid-inducible system from pseudomonas putida for orthogonal gene expression control in escherichia coli and cupriavidus necator |
| url | https://eprints.nottingham.ac.uk/42195/ https://eprints.nottingham.ac.uk/42195/ https://eprints.nottingham.ac.uk/42195/ |