3 years ago

Genetic Encoding of Photocaged Tyrosines with Improved Light-Activation Properties for the Optical Control of Protease Function

Genetic Encoding of Photocaged Tyrosines with Improved Light-Activation Properties for the Optical Control of Protease Function
Alexander Deiters, Jessica Torres-Kolbus, Ji Luo, Jihe Liu
We genetically encoded three new caged tyrosine analogues with improved photochemical properties by using an engineered pyrrolysyl-tRNA synthetase/tRNACUA pair in bacterial and mammalian cells. We applied the new tyrosine analogues to the photoregulation of firefly luciferase by caging its key tyrosine residue, Tyr340, and observed excellent off-to-on light switching. This reporter was then used to evaluate the activation rates of the different light-removable protecting groups in live cells. We identified the nitropiperonyl caging group as an excellent compromise between incorporation efficiency and photoactivation properties. To demonstrate applicability of the new caged tyrosines, an important proteolytic enzyme, tobacco etch virus (TEV) protease, was engineered for optical control. The ability to incorporate differently caged tyrosine analogues into proteins in live cells further expands the unnatural amino acid and optogenetic toolbox. Turn on the lights! Three new caged tyrosine analogues with improved light-activation properties were genetically encoded in bacterial and mammalian cells. The caged tyrosines were applied to the optical control of firefly luciferase and tobacco etch virus (TEV) protease, and excellent off-to-on light switching was observed.

Publisher URL: http://onlinelibrary.wiley.com/resolve/doi

DOI: 10.1002/cbic.201700147

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