Dynamics and mechanism of off- to on-switching in Dreiklang a decoupled reversibly switchable fluorescent protein

Fatima, Anam, He, Yongle, Rosenberger, Danielle, Greetham, Gregory M., Malakar, Partha, Lukacs, Andras, Tonge, Peter J. and Meech, Stephen R. (2025) Dynamics and mechanism of off- to on-switching in Dreiklang a decoupled reversibly switchable fluorescent protein. Angewandte Chemie International Edition. ISSN 1433-7851

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Abstract

Dreiklang is a reversibly switchable (rs) fluorescent protein (FP) with a unique off-state, a UV absorbing hydrated form of the typical FP chromophore. Here we report ultrafast dynamics of the off- to on-state transition in Dreiklang using complementary ultrafast optical and vibrational transient absorption to resolve chromophore driven protein structural dynamics. This approach allows observation of the real-time response in a protein to bond breaking and forming events. The excited electronic state decays in a nonsingle exponential fashion in tens to hundreds of picoseconds, undergoing photodehydration with a yield of several per-cent. The primary photoproduct formed is identified as the cis protonated form of the FP chromophore, initially in a perturbed H-bonded environment. This primary product relaxes on a few microseconds timescale by a mechanism involving changes to a glutamic acid residue and modifications of the amide backbone, possibly involving a carbonyl to imine tautomerization. The temporal and spectral resolution of Dreiklang's photodehydration provides data against which to test quantum chemical calculations of reaction dynamics in proteins and suggests a route to modifying and potentially enhancing its photoswitching properties.

Item Type:	Article
Additional Information:	Data Availability Statement: The data that support the findings of this study are available from the corresponding author upon reasonable request.
Faculty \ School:	Faculty of Science > School of Chemistry, Pharmacy and Pharmacology
UEA Research Groups:	Faculty of Science > Research Groups > Centre for Photonics and Quantum Science Faculty of Science > Research Groups > Chemistry of Light and Energy
Related URLs:	https://onlinelibrary.wiley.com/doi/10.1...
Depositing User:	LivePure Connector
Date Deposited:	14 Nov 2025 16:30
Last Modified:	19 Nov 2025 15:32
URI:	https://ueaeprints.uea.ac.uk/id/eprint/100998
DOI:	10.1002/anie.202518264

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