Engineering ligand-specific biosensors for aromatic amino acids and neurochemicals

• Published in: Cell systems Vol. 13; no. 3; pp. 204 - 214.e4
• Main Authors: Rottinghaus, Austin G., Xi, Chenggang, Amrofell, Matthew B., Yi, Hyojeong, Moon, Tae Seok
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 16.03.2022
• Subjects: Amino Acids, Aromatic; aromatic amino acid; aromatic neurochemical; Biosensing Techniques - methods; directed evolution; Escherichia coli; Ligands; Phenylalanine; probiotic; protein engineering; specific sensors; specific sensors; probiotic; protein engineering; aromatic neurochemical; aromatic amino acid; directed evolution
• ISSN: 2405-4712; 2405-4720
• DOI: 10.1016/j.cels.2021.10.006

Microbial biosensors have diverse applications in metabolic engineering and medicine. Specific and accurate quantification of chemical concentrations allows for adaptive regulation of enzymatic pathways and temporally precise expression of diagnostic reporters. Although biosensors should differentiate structurally similar ligands with distinct biological functions, such specific sensors are rarely found in nature and challenging to create. Using E. coli Nissle 1917, a generally regarded as safe microbe, we characterized two biosensor systems that promiscuously recognize aromatic amino acids or neurochemicals. To improve the sensors’ selectivity and sensitivity, we applied rational protein engineering by identifying and mutagenizing amino acid residues and successfully demonstrated the ligand-specific biosensors for phenylalanine, tyrosine, phenylethylamine, and tyramine. Additionally, our approach revealed insights into the uncharacterized structure of the FeaR regulator, including critical residues in ligand binding. These results lay the groundwork for developing kinetically adaptive microbes for diverse applications. A record of this paper’s transparent peer review process is included in the supplemental information. [Display omitted] •Created specific sensors for structurally similar amino acids and neurochemicals•Developed a protein engineering method requiring only a basic structural understanding•Provided insights into the otherwise uncharacterized structure of a transcription factor This work represents a considerable achievement in protein engineering and includes the ligand-specific sensors for aromatic amino acids such as phenylalanine and tyrosine as well as neurochemicals such as phenylethylamine and tyramine, which are structurally similar and have been implicated in a variety of medical conditions. It lays the groundwork for developing medically relevant probiotic and wearable sensors with high specificity, which is critical to differentiate metabolites with divergent functions and create smart probiotics and cell-free biosensors for accurate diagnostics.