TY - JOUR T1 - Demonstration of <em>de novo</em> chemotaxis in <em>E. coli</em> using a real-time, quantitative, and digital-like approach JF - bioRxiv DO - 10.1101/114207 SP - 114207 AU - Tzila Davidov AU - Naor Granik AU - Sharbel Zahran AU - Inbal Adir AU - Ofek Elul AU - Tal Fried AU - Asif Gil AU - Bar Mayo AU - Shilo Ohayon AU - Shiran Sarig AU - Nofar Shasha AU - Shirane Tsedef AU - Shani Weiner AU - Michal Brunwasser-Meirom AU - Alexandra Ereskovsky AU - Noa Katz AU - Beate Kaufmann AU - Yuri Haimov AU - Heidi Leonard AU - Ester Segal AU - Roee Amit Y1 - 2017/01/01 UR - http://biorxiv.org/content/early/2017/03/06/114207.abstract N2 - Chemotaxis is the movement of an organism in response to an external chemical stimulus. This system enables bacteria to sense their immediate environment and adapt to changes in its chemical composition. Bacterial chemotaxis is mediated by chemoreceptors, membrane proteins that bind an effector and transduce the signal to the downstream proteins. From a synthetic biology perspective, the natural chemotactic repertoire is of little use since bacterial chemoreceptors have evolved to sense specific ligands that either benefit or harm the cell. Here we demonstrate that using a combined computational design approach together with a quantitative, real-time, and digital detection approach, we can rapidly design, manufacture, and characterize a synthetic chemoreceptor in E. coli for histamine (a ligand for which there are no known chemoreceptors). First, we employed a computational protocol that uses the Rosetta bioinformatics software together with high threshold filters to design mutational variants to the native Tar ligand binding domain that target histamine. Second, we tested different ligand-chemoreceptors pairs with a novel chemotaxis assay, based on optical reflectance interferometry of porous silicon (PSi) optical transducers, enabling label-free quantification of chemotaxis by monitoring real-time changes in the optical readout (expressed as the effective optical thickness, EOT). We found that different ligands can be characterized by an individual set of fingerprints in our assay. Namely, a binary, digital-like response in EOT change (i.e. positive or negative) that differentiates between attractants and repellants, the amplitude of change of EOT response, and the rate by which steady state in EOT change is reached. Using this assay, we were able to positively identify and characterize a single mutational chemoreceptor variant for histamine that mediated chemotaxis comparably to the natural Tar-aspartate system. Our results demonstrate the possibility of not only expanding the natural chemotaxis repertoire, but also provide a new quantitative assay by which to characterize the efficacy of the chemotactic response. ER -