A yeast surface display platform for plant hormone receptors: Toward directed evolution of new biosensors |
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| Authors: | Paul J Steiner Matthew A Bedewitz Angélica V Medina-Cucurella Sean R Cutler Timothy A Whitehead |
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| Affiliation: | 1. Department of Chemical and Biological Engineering, University of Colorado, Boulder, Colorado;2. Department of Chemical Engineering and Materials Science, Michigan State University, East Lansing, Michigan;3. Department of Botany and Plant Sciences, University of California, Riverside, California |
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| Abstract: | Small-molecule biosensors have major applications in biotechnology and medicine but remain difficult to engineer. Plant hormone receptors represent an attractive platform for engineering such biosensors because their chemically induced dimerization architectures naturally decouple small-molecule sensing and sensor actuation. Rapid biosensor engineering will require quantitative high-throughput screening methods. Here we develop a yeast surface display (YSD) platform for the PYR1/HAB1 abscisic acid sensor of Arabidopsis thaliana. We extensively optimized PYR1 surface display, HAB1 purification, and binding reaction conditions. Our system reproduces previous results with wild-type and engineered receptors, and a mathematical analysis of the PYR1/HAB1 system allows us to infer all binding constants. Critically, we find that a previously engineered PYR1 receptor with altered ligand specificity binds HAB1 with identical affinity, suggesting that substantial reengineering of the PYR1 binding pocket does not compromise sensor actuation. This YSD platform for A. thaliana PYR1/HAB1 will facilitate future biosensor engineering efforts. |
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| Keywords: | biosensors chemical induced dimerization hormones plant biotechnology yeast surface display |
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