Toward a thermally self-regulating living material

Abstract

Engineered living materials (ELMs) retain desirable characteristics of the living component, such as exponential growth, self-repair, and responsiveness to external stimuli. Escherichia coli are a promising constituent of ELMs because they are very tractable to genetic engineering.Variation in ambient temp. presents a challenge in deploying ELMs outside of a lab. environment. E. coli experience maximal growth near 37°C. In addn., E. coli protein synthesis decreases below 37°C, while protein misfolding and aggregation tends to increase with temp.Here, we develop a genetically encoded mechanism for autonomous temp. homeostasis in ELMs contg. E. coli by engineering circuits that change expression of a light-absorptive chromophore in response to changes in temp. Our simulations show that by increasing absorptivity below 36°C, the material will heat above the ambient temp. to preserve optimal growth and protein expression, and thus material functionality.We program bacteria to respond to temp. using temp.-sensitive transcriptional repressors (TSRs). Two families of TSRs with switching temps. ranging from 36°C to 44°C have been developed in our lab by directed evolution of TcI, a temp.-sensitive mutant of bacteriophage l repressor cI, and TlpA, a transcriptional auto-repressor from the virulence plasmid of Salmonella typhimurium. These thermal bioswitches can be further tuned for optimal switching in the ELM application. Formation of a black chromophore from a pale yellow precursor is enzymically catalyzed. Integrating the gene for this enzyme into a genetic circuit with a down-shifted mutant of TlpA enables E. coli to express black chromophore at low temps. and not at high temps.We measure the ability of patches of E. coli (simulating an E. coli-based ELM) to grow in a custom lighted incubator at different ambient temps. by observing patch diam. and thickness. We continuously monitor E. coli temp. under illumination using thermal IR imaging with custom controller software. Comparison of exptl. results and simulations will be presented. We demonstrate thermal control of pigmentation and resulting increase in sample temp.

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