Abstract
Photosensory protein domains, derived from nature, are foundational for optogenetic protein engineering. Tailoring their properties enables their full exploitation for optogenetic regulation in basic research and applied bioengineering applications. Here, we present a simple, yet powerful strategy based on random mutagenesis coupled to high-throughput screening that allowed altering the most fundamental properties of the widely used nMag/pMag photodimerization system: its light sensitivity and activation. Variants were characterized in vivo in bacteria by flow cytometry and during the entire growth curve by spectrofluorometry. We identify mutations that either increase or decrease the light sensitivity at sub-saturating light intensities, while also improving the light activation and dark-to-light fold change. Notably, light sensitivity and activation levels could be changed independently. In addition, we demonstrated that the shapes of the dose-response curves can be finely tuned. This broadens the applicability of the Magnets photosensors for optogenetic regulation strategies.
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Data availability
Source data are provided with this paper and are available at Zenodo [https://doi.org/10.5281/zenodo.18815874]. Previously published protein structures are available at the PDB under accession code PDB 3RH8. Source data are provided with this paper.
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Acknowledgements
We thank Dr. Tsvetan Kardashliev for helpful discussions and Dr. Luzius Pestalozzi for the testing and the supply of the polymerase and buffer used for error-prone PCR. We further thank Dr. Stephanie Aoki for helpful discussions. We thank the Single Cell and Lab Automation Facility of the DBSSE, ETH Zurich, in particular Dr. Gregor Schmidt, Dr. Aleksandra Gumienny, and Dr. Mariangela Di Tacchio for their excellent support throughout the project. This article is dedicated to the memory of Josep (Pepe) Casadesús. We acknowledge funding from FET-Open research and innovation actions grant under the European Union’s Horizon 2020 research and innovation program (CyGenTiG; grant agreement 801041) to M.K. D.C. was a recipient of an EMBO Short-Term Fellowship (Grant number 8903).
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A.B. conceived, planned, and coordinated the project and wrote the manuscript with contributions from all authors. A.B. and Y.W. generated the libraries and performed the FACS. A.B., Y.W., and D.C. designed and performed bacterial experiments and analyzed the corresponding data. D.C. performed the PC analysis and hierarchical clustering. S.D. performed experiments in mammalian cells and analyzed the corresponding data. M.K. supervised the project and provided funding.
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Baumschlager, A., Weber, Y., Cánovas, D. et al. Enhancing the performance of Magnets photosensors. Nat Commun (2026). https://doi.org/10.1038/s41467-026-70695-7
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DOI: https://doi.org/10.1038/s41467-026-70695-7
