Creation of a point-of-care therapeutics sensor using protein engineering, electrochemical sensing and electronic integration

Point-of-care sensors, which are low-cost and user-friendly, play a crucial role in precision medicine by providing quick results for individuals. Here, we transform the conventional glucometer into a 4-hydroxytamoxifen therapeutic biosensor in which 4-hydroxytamoxifen modulates the electrical signal generated by glucose oxidation. To encode the 4-hydroxytamoxifen signal within glucose oxidation, we introduce the ligand-binding domain of estrogen receptor-alpha into pyrroloquinoline quinone-dependent glucose dehydrogenase by constructing and screening a comprehensive protein insertion library. In addition to obtaining 4-hydroxytamoxifen regulatable engineered proteins, these results unveil the significance of both secondary and quaternary protein structures in propagation of conformational signals. By constructing an effective bioelectrochemical interface, we detect 4-hydroxytamoxifen in human blood samples as changes in the electrical signal and use this to develop an electrochemical algorithm to decode the 4-hydroxytamoxifen signal from glucose. To meet the miniaturization and signal amplification requirements for point-of-care use, we harness power from glucose oxidation to create a self-powered sensor. We also amplify the 4-hydroxytamoxifen signal using an organic electrochemical transistor, resulting in milliampere-level signals. Our work demonstrates a broad interdisciplinary approach to create a biosensor that capitalizes on recent innovations in protein engineering, electrochemical sensing, and electrical engineering.