Batch crystallization of rhodopsin for structural dynamics using an X-ray free-electron laser
Category
Published on
Type
journal-article
Author
Wenting Wu and Przemyslaw Nogly and Jan Rheinberger and Leonhard M. Kick and Cornelius Gati and Garrett Nelson and Xavier Deupi and Jörg Standfuss and Gebhard Schertler and Valérie Panneels
Citation
Wu, W. et al., 2015. Batch crystallization of rhodopsin for structural dynamics using an X-ray free-electron laser. Acta Crystallogr Sect F Struct Biol Cryst Commun, 71(7), pp.856–860. Available at: http://dx.doi.org/10.1107/s2053230x15009966.
Abstract
Rhodopsin is a membrane protein from the G protein-coupled receptor family. Together with its ligand retinal, it forms the visual pigment responsible for night vision. In order to perform ultrafast dynamics studies, a time-resolved serial femtosecond crystallography method is required owing to the nonreversible activation of rhodopsin. In such an approach, microcrystals in suspension are delivered into the X-ray pulses of an X-ray free-electron laser (XFEL) after a precise photoactivation delay. Here, a millilitre batch production of high-density microcrystals was developed by four methodical conversion steps starting from known vapour-diffusion crystallization protocols: (i) screening the low-salt crystallization conditions preferred for serial crystallography by vapour diffusion, (ii) optimization of batch crystallization, (iii) testing the crystal size and quality using second-harmonic generation (SHG) imaging and X-ray powder diffraction and (iv) production of millilitres of rhodopsin crystal suspension in batches for serial crystallography tests; these crystals diffracted at an XFEL at the Linac Coherent Light Source using a liquid-jet setup.
DOI
Funding
NSF-STC Biology with X-ray Lasers (NSF-1231306)
