There is a growing interest in integrating principles from both biology and electronics to enhance molecular communication within living cells. Synthetic biology and electrogenetics pro- vide a framework for utilising redox mediators and electrochemical signals to modulate gene expression in living cells. A redox-sensitive circuit, based on the bacterial stress response system SoxRS, has been studied and implemented. The SoxRS system comprises a transcription factor that undergoes oxidation in the presence of redox mediators, leading to a conformational change enabling recognition of its promoter, pSoxR, and initiation of the biological response. The activation of SoxR is determined by the oxidation state of redox mediators, controllable through electrochemical signals. However, controlling SoxR activation in the presence of oxygen remains challenging due to respiratory interference, limiting its broad adoption by the metabolic engineering community.
This thesis aims to develop and expand a synthetic toolbox based on the electrogenetic SoxR circuit to regulate gene expression using electrochemical signals in aerobic conditions. Firstly, the SoxR redox-sensitive circuit is investigated, assessing its response to various redox cycling molecules, studying its portability to other organisms, and evaluating its impact on cellular metabolism. Secondly, methods for dynamic monitoring of electrogenetic systems are explored, utilising RNA light-up aptamers as transcriptional reporters, unstable fluorescent proteins with high degradation rates, and microfluidic techniques. Thirdly, genetic regulation under aerobic conditions is investigated by implementing a strategy based on the reducing agent TCEP to prevent interference from oxygen in electrochemical regulation. Together, the functionality of the electrogenetic circuit in regulating a metabolic pathway under aerobic conditions is proven, discussing the challenges and showcasing its potential applications in the fields of synthetic biology and metabolic engineering.