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 dynamica... More
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.
