Biosensor libraries harness large classes of binding domains for allosteric transcription regulators

Abstract

Bacteria’s ability to specifically sense small molecules in their environment and trigger metabolic responses in accordance is an invaluable biotechnological resource. While many transcription factors (TFs) mediating these processes have been studied, only a handful has been leveraged for molecular biology applications. To expand this panel of biotechnologically important sensors here we present a strategy for the construction and testing of chimeric TF libraries, based on the fusion of highly soluble periplasmic binding proteins (PBPs) with DNA-binding domains (DBDs). We validated this strategy by constructing and functionally testing two unique sense-and-response regulators for benzoate, an environmentally and industrially relevant metabolite. This work will enable the development of tailored biosensors for synthetic regulatory circuits.

Design of a Transcriptional Biosensor for the Portable, On-Demand Detection of Cyanuric Acid

“…Native promoters also exhibit context dependence, which complicates the porting of aTFs from non-model hosts (e.g., Pseudomonas or Acinetobacter) into common laboratory strains of E. coli. [15][16][17] Thus, repurposing the large diversity of known aTFs and the molecules they sense into useful diagnostic tools requires a better understanding of their fundamental mechanism of action.…”

Chimeric LysR-Type Transcriptional Biosensors for Customizing Ligand Specificity Profiles toward Flavonoids

“…This indicates that, despite the significant similarity of the donor and chassis cir- This is the first contribution of LysR-type chimeric biosensors to the biosensor repertoire which, until now, only contained chimeras from the LacI-and XylR-family. 1,67,68,71,84,85 LysRtype regulators comprise the largest known family of prokaryotic TFs and are able to detect a wide variety of molecules such as arginine, acetic acid, malonate, salicylates, chitobiose and flavonoids. 62,86 Therefore, the proposed strategies could also be used to generate various other chimeric LysR-type biosensors for different ligand molecules, whether or not within…”

Advanced strategies and tools to facilitate and streamline microbial adaptive laboratory evolution

“…First, new genetic biosensors can be identified through transcriptomic and proteomic analyses of natural organisms [131,132]. Further, toggled screening of mutant libraries (mutated from existing genetic biosensors or generated by the artificial fusion of diverse ligand-binding proteins with DNA-binding domains) by FACS is also a practical and valid method [133][134][135]. Moreover, some progress has been made on their de novo computational design [136,137].…”