Dataset Information

Production, Purification and Characterization of Recombinant Biotinylated Phytochrome B for Extracellular Optogenetics.

ABSTRACT: In the field of extracellular optogenetics, photoreceptors are applied outside of cells to obtain systems with a desired functionality. Among the diverse applied photoreceptors, phytochromes are the only ones that can be actively and reversibly switched between the active and inactive photostate by the illumination with cell-compatible red and far-red light. In this protocol, we describe the production of a biotinylated variant of the photosensory domain of A. thaliana phytochrome B (PhyB-AviTag) in E. coli with a single, optimized expression plasmid. We give detailed instructions for the purification of the protein by immobilized metal affinity chromatography and the characterization of the protein in terms of purity, biotinylation, spectral photoswitching and the light-dependent interaction with its interaction partner PIF6. In comparison to previous studies applying PhyB-AviTag, the optimized expression plasmid used in this protocol simplifies the production process and shows an increased yield and purity.

SUBMITTER: Horner M

PROVIDER: S-EPMC7842835 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

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Production, Purification and Characterization of Recombinant Biotinylated Phytochrome B for Extracellular Optogenetics.

Hörner Maximilian M Yousefi O Sascha OS Schamel Wolfgang W A WWA Weber Wilfried W

Bio-protocol 20200305 5

In the field of extracellular optogenetics, photoreceptors are applied outside of cells to obtain systems with a desired functionality. Among the diverse applied photoreceptors, phytochromes are the only ones that can be actively and reversibly switched between the active and inactive photostate by the illumination with cell-compatible red and far-red light. In this protocol, we describe the production of a biotinylated variant of the photosensory domain of <i>A. thaliana</i> phytochrome B (PhyB ...[more]

PMID: 33659515

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Dataset Information

Production, Purification and Characterization of Recombinant Biotinylated Phytochrome B for Extracellular Optogenetics.

Publications

Production, Purification and Characterization of Recombinant Biotinylated Phytochrome B for Extracellular Optogenetics.

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