Chromophore twisting in the excited state of a photoswitchable fluorescent protein captured by time-resolved serial femtosecond crystallography
Nicolas Coquelle
(1)
,
Michel Sliwa
(2)
,
Joyce Woodhouse
(1)
,
Giorgio Schirò
(1)
,
Virgile Adam
(1)
,
Andrew Aquila
(3)
,
Thomas R. M. Barends
(4)
,
Sébastien Boutet
(3)
,
Martin Byrdin
(1)
,
Sergio Carbajo
(3)
,
Eugenio de La Mora
(1)
,
R. Bruce Doak
(4)
,
Mikolaj Feliks
(3)
,
Franck Fieschi
(1)
,
Lutz Foucar
(4)
,
Virginia Guillon
(1)
,
Mario Hilpert
(4)
,
Mark S. Hunter
(3)
,
Stefan Jakobs
(5)
,
Jason E. Koglin
(3)
,
Gabriela Kovacsova
(4)
,
Thomas J. Lane
(3)
,
Bernard C. Levy
(6)
,
Mengning Liang
(3)
,
Karol Nass
(4)
,
Jacqueline Ridard
(6)
,
Joseph S. Robinson
(3)
,
Christopher M. Roome
(4)
,
Cyril Ruckebusch
(2)
,
Matthew Seaberg
(3)
,
Michel Thépaut
(1)
,
Marco Cammarata
(7)
,
Isabelle Demachy
(6)
,
Martin J Field
(1)
,
Robert L. Shoeman
(8)
,
Dominique Bourgeois
(1)
,
Jacques-Philippe Colletier
(1)
,
Ilme Schlichting
(4)
,
Martin Weik
(1)
1
IBS - UMR 5075 -
Institut de biologie structurale
2 LASIRE - Laboratoire Avancé de Spectroscopie pour les Intéractions la Réactivité et l'Environnement - UMR 8516
3 LCLS - Linac Coherent Light Source
4 Max Planck Institute for Medical Research [Heidelberg]
5 Department of NanoBiophotonics [Göttingen]
6 LCPO - Laboratoire de Chimie Physique D'Orsay
7 IPR - Institut de Physique de Rennes
8 Max-Planck-Institut für Medizinische Forschung
2 LASIRE - Laboratoire Avancé de Spectroscopie pour les Intéractions la Réactivité et l'Environnement - UMR 8516
3 LCLS - Linac Coherent Light Source
4 Max Planck Institute for Medical Research [Heidelberg]
5 Department of NanoBiophotonics [Göttingen]
6 LCPO - Laboratoire de Chimie Physique D'Orsay
7 IPR - Institut de Physique de Rennes
8 Max-Planck-Institut für Medizinische Forschung
Michel Sliwa
- Function : Author
- PersonId : 17467
- IdHAL : michel-sliwa
- ORCID : 0000-0002-5073-8180
- IdRef : 08084166X
Giorgio Schirò
- Function : Author
- PersonId : 742723
- IdHAL : giorgio-schiro
Virgile Adam
- Function : Author
- PersonId : 4048
- IdHAL : virgile-adam
- ORCID : 0000-0003-2209-7846
- IdRef : 250406187
Martin Byrdin
- Function : Author
- PersonId : 1638
- IdHAL : martin-byrdin
- ORCID : 0000-0002-6389-8714
- IdRef : 185869890
Franck Fieschi
- Function : Author
- PersonId : 761070
- ORCID : 0000-0003-1194-8107
- IdRef : 087253240
Karol Nass
- Function : Author
- PersonId : 773540
- ORCID : 0000-0001-8843-1566
Cyril Ruckebusch
- Function : Author
- PersonId : 847898
Marco Cammarata
- Function : Author
- PersonId : 491
- IdHAL : marco-cammarata
- ORCID : 0000-0003-3013-1186
- IdRef : 229610080
Dominique Bourgeois
- Function : Author
- PersonId : 1634
- IdHAL : dominique-bourgeois
- ORCID : 0000-0002-1862-7712
- IdRef : 079965415
Jacques-Philippe Colletier
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- Function : Correspondent author
- PersonId : 739662
- IdHAL : jacques-philippe-colletier
- ORCID : 0000-0003-1819-4686
- IdRef : 110264983
Connectez-vous pour contacter l'auteur
Abstract
Chromophores absorb light in photosensitive proteins and thereby initiate fundamental biological processes such as photosynthesis, vision and biofluorescence. An important goal in their understanding is the provision of detailed structural descriptions of the ultrafast photochemical events that they undergo, in particular of the excited states that connect chemistry to biological function. Here we report on the structures of two excited states in the reversibly photoswitchable fluorescent protein rsEGFP2. We populated the states through femtosecond illumination of rsEGFP2 in its non-fluorescent off state and observed their build-up (within less than one picosecond) and decay (on the several picosecond timescale). Using an X-ray free-electron laser, we performed picosecond time-resolved crystallography and show that the hydroxybenzylidene imidazolinone chromophore in one of the excited states assumes a near-canonical twisted configuration halfway between the trans and cis isomers. This is in line with excited-state quantum mechanics/molecular mechanics and classical molecular dynamics simulations. Our new understanding of the structure around the twisted chromophore enabled the design of a mutant that displays a twofold increase in its off-to-on photoswitching quantum yield.