Here we introduce Z-lock, an optogenetic approach for reversible, light-controlled steric inhibition of protein active sites. The light oxygen voltage (LOV) domain and Zdk, a small protein that binds LOV selectively in the dark, are appended to the protein of interest where they sterically block the active site. Irradiation causes LOV to change conformation and release Zdk, exposing the active site. Computer-assisted protein design was used to optimize linkers and Zdk-LOV affinity, for both effective binding in the dark, and effective light-induced release of the intramolecular interaction. Z-lock cofilin was shown to have actin severing ability in vitro, and in living cancer cells it produced protrusions and invadopodia. An active fragment of the tubulin acetylase αTAT was similarly modified and shown to acetylate tubulin on irradiation.
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The data that support the findings of this study are available from the corresponding authors upon request.
Code is available from the authors upon request or at http://www.hahnlab.com.
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We thank the NIH for supporting this work (grant no. GM122596 to K.M.H., grant nos. CA150344 and CA216248 to J.S.C.). O.J.S. is a recipient of a Ruth L. Kirschstein National Research Service Award (no. 1F31CA192739). We thank M. Azoitei for assisting with Rosetta modeling, T. Watanabe for assistance with localized photoactivation experiments and C. Onyeji for help with cloning and biochemical assays. N.P. was supported by grant PF-16-18-01-CSM from the American Cancer Society.
The authors declare no competing interests.
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Stone, O.J., Pankow, N., Liu, B. et al. Optogenetic control of cofilin and αTAT in living cells using Z-lock. Nat Chem Biol 15, 1183–1190 (2019). https://doi.org/10.1038/s41589-019-0405-4
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