Engineering bacterial motility towards hydrogen-peroxide

Synthetic biologists construct innovative genetic/biological systems to treat environmental, energy, and health problems. Many systems employ rewired cells for non-native product synthesis, while a few have employed the rewired cells as ‘smart’ devices with programmable function. Building on the latter, we developed a genetic construct to control and direct bacterial motility towards hydrogen peroxide, one of the body’s immune response signaling molecules. A motivation for this work is the creation of cells that can target and autonomously treat disease, the latter signaled by hydrogen peroxide release. Bacteria naturally move towards a variety of molecular cues (e.g., nutrients) in the process of chemotaxis. In this work, we engineered bacteria to recognize and move towards hydrogen peroxide, a non-native chemoattractant and potential toxin. Our system exploits oxyRS, the native oxidative stress regulon of E. coli. We first demonstrated H(2)O(2)-mediated upregulation motility regulator, CheZ. Using transwell assays, we showed a two-fold increase in net motility towards H(2)O(2). Then, using a 2D cell tracking system, we quantified bacterial motility descriptors including velocity, % running (of tumble/run motions), and a dynamic net directionality towards the molecular cue. In CheZ mutants, we found that increased H(2)O(2) concentration (0–200 μM) and induction time resulted in increased running speeds, ultimately reaching the native E. coli wild-type speed of ~22 μm/s with a ~45–65% ratio of running to tumbling. Finally, using a microfluidic device with stable H(2)O(2) gradients, we characterized responses and the potential for “programmed” directionality towards H(2)O(2) in quiescent fluids. Overall, the synthetic biology framework and tracking analysis in this work will provide a framework for investigating controlled motility of E. coli and other ‘smart’ probiotics for signal-directed treatment.