Modular Fluorescent Carbon Nanotube Sensors for Discovery and Optimization of Cell Free Expressed Peptides and Enzymes

High-throughput screening and development of new biological molecules from cell free reactions requires modular sensors that may be deployed in vitro with minimal preparation and few interactions with a complex background. As a result of their distinctly strong surface sensitivity, semiconducting single-walled carbon nanotubes (SWNT) are a promising fluorescent transducer for such applications. SWNT can be rendered sensitive to target molecules by non-covalent functionalization with a minimalist binding “corona” that functions without the relatively complex energy donor-acceptor structure typical of colloidal sensors. While this corona may to be used to bind and detect specific analytes, it may be further developed to mimic biological elements and transduce relevant phenomena such as degradation or modification. Herein, we demonstrate fluorescent SWNT sensors which transduce perturbations to selected, biological coronas. By suspending SWNT with enzyme substrates such as proteins, polysaccharides, oligosaccharides, polyphenols, or lipids, SWNT may be rendered sensitive to hydrolases and oxidoreductases. By suspending SWNT with amphiphilic phospholipids or lipopolysaccharides, this corona may simulate the outer surface of a microbe which may be used as a model to screen new cell penetrating therapeutics or antimicrobials. This sensor’s simplicity is promising for microfluidics, where SWNT sensors may be used to substitute inconsistent reagents or complex assays. By further coupling this tool with microfluidic CFPS, this tool may be used for guided evolution of proteins and small peptides for industrial and biomedical applications.