Method for developing optical sensors using a synthetic dye fluorescent protein FRET pair and computational modeling and assessment

book chapter Method for developing optical sensors using a synthetic dye fluorescent protein FRET pair and computational modeling and assessment Methods in Molecular Biology published yes Joshua Mitchell author William Zhang author Michel Herde author Christian Henneberger author Harald L Janovjak author 33BA6C30-F248-11E8-B48F-1D18A9856A870000-0002-8023-9315 Megan O'Mara author Colin Jackson author ViktorStein editor HaJa department Biosensors that exploit Forster resonance energy transfer (FRET) can be used to visualize biological and physiological processes and are capable of providing detailed information in both spatial and temporal dimensions. In a FRET-based biosensor, substrate binding is associated with a change in the relative positions of two fluorophores, leading to a change in FRET efficiency that may be observed in the fluorescence spectrum. As a result, their design requires a ligand-binding protein that exhibits a conformational change upon binding. However, not all ligand-binding proteins produce responsive sensors upon conjugation to fluorescent proteins or dyes, and identifying the optimum locations for the fluorophores often involves labor-intensive iterative design or high-throughput screening. Combining the genetic fusion of a fluorescent protein to the ligand-binding protein with site-specific covalent attachment of a fluorescent dye can allow fine control over the positions of the two fluorophores, allowing the construction of very sensitive sensors. This relies upon the accurate prediction of the locations of the two fluorophores in bound and unbound states. In this chapter, we describe a method for computational identification of dye-attachment sites that allows the use of cysteine modification to attach synthetic dyes that can be paired with a fluorescent protein for the purposes of creating FRET sensors. Springer2017 eng Synthetic Protein Switches 1064-374510.1007/978-1-4939-6940-1_6 159689 - 99 Mitchell J, Zhang W, Herde M, et al. Method for developing optical sensors using a synthetic dye fluorescent protein FRET pair and computational modeling and assessment. In: Stein V, ed. Synthetic Protein Switches. Vol 1596. Synthetic Protein Switches. Springer; 2017:89-99. doi:<a href="https://doi.org/10.1007/978-1-4939-6940-1_6">10.1007/978-1-4939-6940-1_6</a> Mitchell J, Zhang W, Herde M, Henneberger C, Janovjak HL, O’Mara M, Jackson C. 2017.Method for developing optical sensors using a synthetic dye fluorescent protein FRET pair and computational modeling and assessment. In: Synthetic Protein Switches. Methods in Molecular Biology, vol. 1596, 89–99. J. Mitchell, W. Zhang, M. Herde, C. Henneberger, H.L. Janovjak, M. O’Mara, C. Jackson, in:, V. Stein (Ed.), Synthetic Protein Switches, Springer, 2017, pp. 89–99. J. Mitchell et al., “Method for developing optical sensors using a synthetic dye fluorescent protein FRET pair and computational modeling and assessment,” in Synthetic Protein Switches, vol. 1596, V. Stein, Ed. Springer, 2017, pp. 89–99. Mitchell, Joshua, et al. “Method for Developing Optical Sensors Using a Synthetic Dye Fluorescent Protein FRET Pair and Computational Modeling and Assessment.” Synthetic Protein Switches, edited by Viktor Stein, vol. 1596, Springer, 2017, pp. 89–99, doi:<a href="https://doi.org/10.1007/978-1-4939-6940-1_6">10.1007/978-1-4939-6940-1_6</a>. Mitchell, J., Zhang, W., Herde, M., Henneberger, C., Janovjak, H. L., O’Mara, M., & Jackson, C. (2017). Method for developing optical sensors using a synthetic dye fluorescent protein FRET pair and computational modeling and assessment. In V. Stein (Ed.), Synthetic Protein Switches (Vol. 1596, pp. 89–99). Springer. <a href="https://doi.org/10.1007/978-1-4939-6940-1_6">https://doi.org/10.1007/978-1-4939-6940-1_6</a> Mitchell, Joshua, William Zhang, Michel Herde, Christian Henneberger, Harald L Janovjak, Megan O’Mara, and Colin Jackson. “Method for Developing Optical Sensors Using a Synthetic Dye Fluorescent Protein FRET Pair and Computational Modeling and Assessment.” In Synthetic Protein Switches, edited by Viktor Stein, 1596:89–99. Synthetic Protein Switches. Springer, 2017. <a href="https://doi.org/10.1007/978-1-4939-6940-1_6">https://doi.org/10.1007/978-1-4939-6940-1_6</a>. 9582018-12-11T11:49:24Z2025-07-10T12:01:54Z