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Title: Batch crystallization of rhodopsin for structural dynamics using an X-ray free-electron laser

A new batch preparation method is presented for high-density micrometre-sized crystals of the G protein-coupled receptor rhodopsin for use in time-resolved serial femtosecond crystallography at an X-ray free-electron laser using a liquid jet. 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.
Authors:
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  1. Paul Scherrer Institute, OFLC/103, 5232 Villigen-PSI (Switzerland)
Publication Date:
OSTI Identifier:
22389083
Resource Type:
Journal Article
Resource Relation:
Journal Name: Acta crystallographica. Section F, Structural biology communications; Journal Volume: 71; Journal Issue: Pt 7; Other Information: PMCID: PMC4498706; PMID: 26144230; PUBLISHER-ID: ic5093; PUBLISHER-ID: S2053230X15009966; OAI: oai:pubmedcentral.nih.gov:4498706; Copyright (c) Wu et al. 2015; This is an open-access article distributed under the terms of the Creative Commons Attribution Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original authors and source are cited.; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United States
Language:
English
Subject:
75 CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND SUPERFLUIDITY; CRYSTALLIZATION; CRYSTALLOGRAPHY; CRYSTALS; DENSITY; DIFFUSION; DYNAMICS; ELECTRONS; HARMONIC GENERATION; JETS; LIGANDS; LIGHT SOURCES; LIQUIDS; PULSES; SCREENING