Membrane protein megahertz crystallography at the European XFEL

The world’s first superconducting megahertz repetition rate hard X-ray free-electron laser (XFEL), the European XFEL, began operation in 2017, featuring a unique pulse train structure with 886 ns between pulses. With its rapid pulse rate, the European XFEL may alleviate some of the increasing demand for XFEL beamtime, particularly for membrane protein serial femtosecond crystallography (SFX), leveraging orders-of-magnitude faster data collection. Here, we report the first membrane protein megahertz SFX experiment, where we determined a 2.9 Å-resolution SFX structure of the large membrane protein complex, Photosystem I, a > 1 MDa complex containing 36 protein subunits and 381 cofactors. We address challenges to megahertz SFX for membrane protein complexes, including growth of large quantities of crystals and the large molecular and unit cell size that influence data collection and analysis. The results imply that megahertz crystallography could have an important impact on structure determination of large protein complexes with XFELs.

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004: Informatik

530: Physik

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We acknowledge funding from the Biodesign Center for Applied Structural Discovery at Arizona State University, and the following federal grants: the National Science Foundation (NSF) Science awards for Technology Center (STC) BioXFEL award no. STC-1231306 and award no. 1565180 (N.A.Z., R.A.K., S.B., and J.C.H.S.), the U.S. Department of Energy, Office of Science, Basic Energy Sciences award DE-SC0002164 and DESC0010575, and the National Institutes of Health grant R01GM095583. This work is also supported by the AXSIS project funded by the European Research Council under the European Union Seventh Framework Program (FP/2007–2013)/ERC Grant Agreement no. 609920. Funding was provided by the excellence cluster The Hamburg Center for Ultrafast Imaging—Structure, Dynamics, and Control of Matter at the Atomic Scale of the Deutsche Forschungsgemeinschaft (CUI, DFG-EXC1074), and the BMBF through the Roentgen-Angstrom Cluster grant 05K18CHA. This work was performed, in part, under the auspices of the U.S. Department of Energy by Lawrence Livermore National Laboratory under Contract DE-AC52-07NA27344. M.A.C,, M.L.S., and M.F. were supported by NIH grant R01GM117342. We acknowledge the support of the Australian Research Council through the Centre of Excellence in Advanced Molecular Imaging (CE140100011).

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Membrane protein megahertz crystallography at the European XFEL