Incorporation of bacterial quorum sensing in synthetic biology

Title of Dissertation: INCORPORATION OF BACTERIAL QUORUM SENSING IN SYNTHETIC BIOLOGY Hsuan-Chen Wu, Doctor of Philosophy, 2012 Directed By: Professor William E. Bentley Fischell Department of Bioengineering The global objective of this research is to develop a synthetic biology toolkit consisting of molecules, cells, and devices that provide flexible, yet selective targeting, sensing, and switching capabilities, that in turn guide biological behavior in userspecified manner. We employ bacteria as “smart” programmable devices. We envision creating bacteria that autonomously move to specific areas, synthesize a drug, deliver the drug, and move on to new sites. “Targeting” endows bacterial cells the means to dock onto specific surfaces with antibody-antigen specificity. Sensing and switching capability allows bacteria to sense and, after making a “decision”, respond by synthesizing and delivering cargo to molecular scale features displayed on target surfaces. Relevant surface features may include an overexpressed receptor on a tumor cell, glucagon-like peptide-1 receptor on pancreatic beta cells, or even other bacterial cells resident in a recalcitrant biofilm. Towards the realization of this goal, we employed an antibody-binding protein G display strategy to complex target-binding antibodies with bacteria. We characterized the assembly and efficacy of this complex by binding to well-defined surfaces decorated with specific antigens. For sensing and switching we made use of the genetic circuitry of bacterial quorum sensing (QS) that coordinates multicellular responses. In particular, we hypothesized the creation of a biological “switch” that would take action only after a certain threshold “feature” density had been detected. Specifically, in our most significant demonstration we designed and implemented QS based sensing and actuating based on the surface density of cancer-indicating EGFR receptors displayed on epithelial

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