Assessment of Colorimetric Reporter Enzymes in the PURE System

• Published in: ACS synthetic biology 2021-11, Vol.10 (11), p.3205-3208
• Main Authors: Sharpes, Caitlin E, McManus, John B, Blum, Steven M, Mgboji, Glory E, Lux, Matthew W
• Format: Article
• Language: English
• Subjects: Biosensing Techniques - methods; Cell-Free System - metabolism; Color; Colorimetry - methods; Enzymes - metabolism; Gene Expression - physiology
• ISSN: 2161-5063; 2161-5063
• DOI: 10.1021/acssynbio.1c00360

Colorimetric reporter enzymes are useful for generating eye-readable biosensor readouts that do not require a device to interpret, an attractive property for applications in remote or developing parts of the world. The use of cell-free gene expression further facilitates such applications via amenability to lyophilization and incorporation into materials like paper. Currently, detection of multiple analytes simultaneously with these systems requires multiple reactions or a device. Here we evaluate seven enzymes and 15 corresponding substrates for functionality in a particular cell-free expression system known as PURE. We report eight enzyme/substrate pairs spanning four enzymes that are compatible with PURE. Of the four enzymes, three pairings exhibit no cross-reactivity. We finally show that at least one pairing can be used to create a third color when both are present, highlighting the potential use of these reporters for multiplex sensing.