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Optical Control of Transcription: Genetically Encoded Photoswitchable Variants of T7 RNA Polymerase
Author(s) -
Seifert Swantje,
Ehrt Christiane,
Lückfeldt Lena,
Lubeck Melissa,
Schramm Frederik,
Brakmann Susanne
Publication year - 2019
Publication title -
chembiochem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.201900298
Subject(s) - cryptochrome , t7 rna polymerase , allosteric regulation , transcription (linguistics) , fusion protein , polymerase , computational biology , biology , chemistry , biophysics , microbiology and biotechnology , biochemistry , enzyme , gene , circadian clock , escherichia coli , bacteriophage , recombinant dna , linguistics , philosophy
Light‐sensing protein domains that link an exogenous light signal to the activity of an enzyme have attracted much attention for the engineering of new regulatory mechanisms into proteins and for studying the dynamic behavior of intracellular reactions and reaction cascades. Light–oxygen–voltage (LOV) photoreceptors are blue‐light‐sensing modules that have been intensely characterized for this purpose and linked to several proteins of interest. For the successful application of these tools, it is crucial to identify appropriate fusion strategies for combining sensor and enzyme domains that sustain activity and light‐induced responsivity. Terminal fusion of LOV domains is the natural strategy; however, this is not transferrable to T7 RNA polymerase because both of its termini are involved in catalysis. It is shown herein that it is possible to covalently insert LOV domains into the polymerase protein, while preserving its activity and generating new light‐responsive allosteric coupling.