Photoactivatable Rac1: a photosensor tilt by time-resolved crystallography and a partially open switch I when GDP-bound

Pachota, M.; Shankar, M. K.; Wisniewska, M.; Koczurowska, A.; Casadei, C. M.; Solecka, A.; Kapetanaki, S. M.; Olasz, B.; Lopes Cavalcante, L.; Ke, R.; Lang, D.; Bjelcic, M.; Asghar, A.; Milas, M.; Ursby, T.; Chenchiliyan, M.; Nan, J.; Rosta, E.; Nogly, P.

doi:10.64898/2026.10.27.762334

bioRxiv · 10.64898/2026.10.27.762334

Photoactivatable Rac1: a photosensor tilt by time-resolved crystallography and a partially open switch I when GDP-bound

Pachota, M.·Shankar, M. K.·Wisniewska, M.·Koczurowska, A.·Casadei, C. M.·Solecka, A.·Kapetanaki, S. M.·Olasz, B.·Lopes Cavalcante, L.·Ke, R.·Lang, D.·Bjelcic, M.·Asghar, A.·Milas, M.·Ursby, T.·Chenchiliyan, M.·Nan, J.·Rosta, E.·Nogly, P.

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Abstract

The LOV2 domain of Avena sativa phototropin 1 is widely used to control proteins with light, but how its signal reaches a fused effector has not been visualized. We followed photoactivatable Rac1 by time-resolved serial crystallography at room temperature. Within 350 {micro}s, a fifth of the photosensors form the flavin-cysteine adduct and tilt as a rigid body on Rac1 about an axis through the domain interface, while the J helix stays docked and Rac1 keeps GTP and dark-state switches up to 1.85 ms. Dark crystals contain a variable GDP-bound sub-population, so light data must be referenced to interleaved dark frames. In this sub-population, switch I is only partially open, held at a Ca2+ ion that bridges LOV2 and Rac1, a conformation that can persist in simulations without the lattice. The photosensor thus first acts on Rac1 at the interface that also holds switch I in the GDP-bound state.

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Pachota, M., Shankar, M. K., Wisniewska, M., Koczurowska, A., Casadei, C. M., Solecka, A., Kapetanaki, S. M., Olasz, B., Lopes Cavalcante, L., Ke, R., Lang, D., Bjelcic, M., Asghar, A., Milas, M., Ursby, T., Chenchiliyan, M., Nan, J., Rosta, E., Nogly, P.. 2026-10-28. Photoactivatable Rac1: a photosensor tilt by time-resolved crystallography and a partially open switch I when GDP-bound. https://doi.org/10.64898/2026.10.27.762334

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