Synthetic Biology Combined With Natural Biochemical Pathways For Engineering Robust Whole-Cell Biosensors

To date, the main guideline in whole-cell biosensors engineering has mainly based on fusion of a single stress-responsive promoter to a reporter protein within living cell. However, such concept has greatly failed to deal with real-world samples which require higher computational complexity, specificity and stability. Bacterial biosensors implement rapid and cheap monitoring of the presence of toxicants in water, air, soil and food by translating the concentration of a specific analyte real-time into a measurable signal. Biosensor engineering involves designing genetic circuits, which lie in the basis of the rapidly evolving discipline of synthetic biology, thus enabling reengineering and rewiring natural biological pathways for both the discovery of the principles lying in the basis of regulatory networks and practical applications in areas such as medicine, environmental bioremediation, biofuels manufacturing and biological computation. In bacterial biosensors, the stress response is usually linked to complex cascades involving broad and global regulatory circuits, generating a semispecific reaction. To overcome these challenges, we propose to combine natural biochemical networks together with strategies implemented in the framework of synthetic biology to build robust whole-cell biosensors. To increase the specificity of bacterial biosensors, the stress promoters were combined with synthetic promoters that are regulated by a repressor/activator protein sensitive only to a specific compound. To reduce the basal level activity as well as to receive a high separation between the maximum and minimum activity we incorporated a negative feedback loop. Finally, to adjust the response between different stress-promoters we used signal amplification design.