Development of a Genetically Encoded Biosensor for Detection of Polyketide Synthase Extender Units in Escherichia coli
| Autor: | Gavin J. Williams, Edward Kalkreuter, Aaron M Keeler, Anuran K Gayen, Kyle S Bingham, Alexandra A Malico |
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| Rok vydání: | 2019 |
| Předmět: |
0106 biological sciences
Biomedical Engineering Computational biology macromolecular substances Biosensing Techniques medicine.disease_cause Protein Engineering 01 natural sciences Biochemistry Genetics and Molecular Biology (miscellaneous) Article 03 medical and health sciences chemistry.chemical_compound Synthetic biology Polyketide Biosynthesis 010608 biotechnology Polyketide synthase medicine Escherichia coli 030304 developmental biology 0303 health sciences biology Effector Escherichia coli Proteins General Medicine Recombinant Proteins Malonyl Coenzyme A Malonyl-CoA chemistry Polyketides biology.protein lipids (amino acids peptides and proteins) Synthetic Biology Biosensor Polyketide Synthases Metabolic Networks and Pathways Transcription Factors |
| Zdroj: | ACS Synth Biol |
| ISSN: | 2161-5063 |
| Popis: | The scaffolds of polyketides are constructed via assembly of extender units based on malonyl-CoA and its derivatives that are substituted at the C2-position with diverse chemical functionality. Subsequently, a transcription factor-based biosensor for malonyl-CoA has proven to be a powerful tool for detecting malonyl-CoA, facilitating the dynamic regulation of malonyl-CoA biosynthesis, and guiding high-throughput engineering of malonyl-CoA dependent processes. Yet, a biosensor for the detection of malonyl-CoA derivatives has yet to be reported, severely restricting the application of high-throughput synthetic biology approaches to engineering extender unit biosynthesis and limiting the ability to dynamically regulate the biosynthesis of polyketide products that are dependent on such α-carboxyacyl-CoAs. Herein, the FapR biosensor was re-engineered and optimized for a range of mCoA concentrations across a panel of E. coli strains. The effector specificity of FapR was probed by cell-free transcription-translation, revealing that a variety of non-native and non-natural acyl-thioesters are FapR effectors. This FapR promiscuity proved sufficient for the detection of the polyketide extender unit methylmalonyl-CoA in E. coli, providing the first reported genetically encoded biosensor for this important metabolite. As such, the previously unknown broad effector promiscuity of FapR provides a platform to develop new tools and approaches that can be leveraged to overcome limitations of pathways that construct diverse α-carboxyacyl-CoAs and those that are dependent on them, including biofuels, antibiotics, anticancer drugs, and other value-added products. |
| Databáze: | OpenAIRE |
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