CL6mN: Rationally Designed Optogenetic Photoswitches with Tunable Dissociation DynamicsClick to copy article linkArticle link copied!
- Abhirup MukherjeeAbhirup MukherjeeDepartment of Chemical and Biomolecular Engineering, Georgia Institute of Technology, Atlanta, Georgia 30332, United StatesMore by Abhirup Mukherjee
- Chaitanya SudrikChaitanya SudrikDepartment of Chemical and Biological Engineering, Rensselaer Polytechnic Institute, Troy, New York 12180, United StatesMore by Chaitanya Sudrik
- Yuge HuYuge HuDepartment of Chemical and Biomolecular Engineering, Georgia Institute of Technology, Atlanta, Georgia 30332, United StatesMore by Yuge Hu
- Manish ArhaManish ArhaCentre for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York 12180, United StatesMore by Manish Arha
- Mark StathosMark StathosBioengineering Graduate Program, Georgia Institute of Technology, Atlanta, Georgia 30332, United StatesMore by Mark Stathos
- Jieung BaekJieung BaekDepartment of Bioengineering, University of California, Berkeley, Berkeley, California 94720, United StatesMore by Jieung Baek
- David V. Schaffer*David V. Schaffer*Tel (510) 643-5963. Fax: 510-642-4778. Email: [email protected]Department of Bioengineering, University of California, Berkeley, Berkeley, California 94720, United StatesDepartment of Chemical Engineering, University of California, Berkeley, Berkeley, California 94720, United StatesHelen Wills Neuroscience Institute, University of California, Berkeley, Berkeley, California 94720, United StatesMore by David V. Schaffer
- Ravi S. Kane*Ravi S. Kane*Tel. (404) 385-4608. Email: [email protected]Department of Chemical and Biomolecular Engineering, Georgia Institute of Technology, Atlanta, Georgia 30332, United StatesMore by Ravi S. Kane
Abstract

The field of optogenetics uses genetically encoded photoswitches to modulate biological phenomena with high spatiotemporal resolution. We report a set of rationally designed optogenetic photoswitches that use the photolyase homology region of A. thaliana cryptochrome 2 (Cry2PHR) as a building block and exhibit highly efficient and tunable clustering in a blue-light dependent manner. CL6mN (Cry2-mCherry-LRP6c with N mutated PPPAP motifs) proteins were designed by mutating and/or truncating five crucial PPP(S/T)P motifs near the C-terminus of the optogenetic Wnt activator Cry2-mCherry-LRP6c, thus eliminating its Wnt activity. Light-induced CL6mN clusters have significantly greater dissociation half-lives than clusters of wild-type Cry2PHR. Moreover, the dissociation half-lives can be tuned by varying the number of PPPAP motifs, with the half-life increasing as much as 6-fold for a variant with five motifs (CL6m5) relative to Cry2PHR. Finally, we demonstrate the compatibility of CL6mN with previously reported Cry2-based photoswitches by optogenetically activating RhoA in mammalian cells.
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This article is cited by 3 publications.
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- Amelia C. McCue, Brian Kuhlman. Design and engineering of light-sensitive protein switches. Current Opinion in Structural Biology 2022, 74 , 102377. https://doi.org/10.1016/j.sbi.2022.102377
- Diego S. Vazquez, Pamela L. Toledo, Alejo R. Gianotti, Mario R. Ermácora. Protein conformation and biomolecular condensates. Current Research in Structural Biology 2022, 4 , 285-307. https://doi.org/10.1016/j.crstbi.2022.09.004
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