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Design, construction, and validation of optogenetic proteins.

Authors
  • O'Banion, Colin P1
  • Goswami, Anwesha1
  • Lawrence, David S2
  • 1 Division of Chemical Biology and Medicinal Chemistry, UNC Eshelman School of Pharmacy, University of North Carolina, Chapel Hill, NC, United States. , (United States)
  • 2 Department of Chemistry, University of North Carolina, Chapel Hill, NC, United States; Department of Pharmacology, University of North Carolina, Chapel Hill, NC, United States. Electronic address: [email protected] , (United States)
Type
Published Article
Journal
Methods in enzymology
Publication Date
Jan 01, 2019
Volume
621
Pages
171–190
Identifiers
DOI: 10.1016/bs.mie.2019.02.019
PMID: 31128778
Source
Medline
Keywords
Language
English
License
Unknown

Abstract

Cellular optogenetics employs light-regulated, genetically encoded protein actuators to perturb cellular signaling with unprecedented spatial and temporal control. Here, we present a potentially generalized approach for transforming a given protein of interest (POI) into an optogenetic species. We describe the rational and methods by which we developed three different optogenetic POIs utilizing the Cry2-Cib photodimerizing pair. The process pipeline is highlighted by (1) developing a low level, constitutively active POI that is independent of endogenous regulation, (2) fusion of the mutant protein of interest to an optogenetic photodimerizing system, and (3) light-mediated recruitment of the light-responsive POI to specific subcellular regions. © 2019 Elsevier Inc. All rights reserved.

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