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Advances in X-ray free electron laser (XFEL) diffraction data processing applied to the crystal structure of the synaptotagmin-1 / SNARE complex.

ABSTRACT: X-ray free electron lasers (XFELs) reduce the effects of radiation damage on macromolecular diffraction data and thereby extend the limiting resolution. Previously, we adapted classical post-refinement techniques to XFEL diffraction data to produce accurate diffraction data sets from a limited number of diffraction images (Uervirojnangkoorn et al., 2015), and went on to use these techniques to obtain a complete data set from crystals of the synaptotagmin-1 / SNARE complex and to determine the structure at 3.5 Å resolution (Zhou et al., 2015). Here, we describe new advances in our methods and present a reprocessed XFEL data set of the synaptotagmin-1 / SNARE complex. The reprocessing produced small improvements in electron density maps and the refined atomic model. The maps also contained more information than those of a lower resolution (4.1 Å) synchrotron data set. Processing a set of simulated XFEL diffraction images revealed that our methods yield accurate data and atomic models.

SUBMITTER: Lyubimov AY

PROVIDER: S-EPMC5094853 | biostudies-literature | 2016 Oct

REPOSITORIES: biostudies-literature

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Publications

Advances in X-ray free electron laser (XFEL) diffraction data processing applied to the crystal structure of the synaptotagmin-1 / SNARE complex.

Lyubimov Artem Y AY Uervirojnangkoorn Monarin M Zeldin Oliver B OB Zhou Qiangjun Q Zhao Minglei M Brewster Aaron S AS Michels-Clark Tara T Holton James M JM Sauter Nicholas K NK Weis William I WI Brunger Axel T AT

eLife 20161012

X-ray free electron lasers (XFELs) reduce the effects of radiation damage on macromolecular diffraction data and thereby extend the limiting resolution. Previously, we adapted classical post-refinement techniques to XFEL diffraction data to produce accurate diffraction data sets from a limited number of diffraction images (Uervirojnangkoorn et al., 2015), and went on to use these techniques to obtain a complete data set from crystals of the synaptotagmin-1 / SNARE complex and to determine the st ...[more]

PMID: 27731796

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Project description:The advent and application of the X-ray free-electron laser (XFEL) has uncovered the structures of proteins that could not previously be solved using traditional crystallography. While this new technology is powerful, optimization of the process is still needed to improve data quality and analysis efficiency. One area is sample heterogeneity, where variations in crystal size (among other factors) lead to the requirement of large data sets (and thus 10-100?mg of protein) for determining accurate structure factors. To decrease sample dispersity, we developed a high-throughput microfluidic sorter operating on the principle of dielectrophoresis, whereby polydisperse particles can be transported into various fluid streams for size fractionation. Using this microsorter, we isolated several milliliters of photosystem I nanocrystal fractions ranging from 200 to 600?nm in size as characterized by dynamic light scattering, nanoparticle tracking, and electron microscopy. Sorted nanocrystals were delivered in a liquid jet via the gas dynamic virtual nozzle into the path of the XFEL at the Linac Coherent Light Source. We obtained diffraction to ?4?Å resolution, indicating that the small crystals were not damaged by the sorting process. We also observed the shape transforms of photosystem I nanocrystals, demonstrating that our device can optimize data collection for the shape transform-based phasing method. Using simulations, we show that narrow crystal size distributions can significantly improve merged data quality in serial crystallography. From this proof-of-concept work, we expect that the automated size-sorting of protein crystals will become an important step for sample production by reducing the amount of protein needed for a high quality final structure and the development of novel phasing methods that exploit inter-Bragg reflection intensities or use variations in beam intensity for radiation damage-induced phasing. This method will also permit an analysis of the dependence of crystal quality on crystal size.

Dataset Information

Advances in X-ray free electron laser (XFEL) diffraction data processing applied to the crystal structure of the synaptotagmin-1 / SNARE complex.

Publications

Advances in X-ray free electron laser (XFEL) diffraction data processing applied to the crystal structure of the synaptotagmin-1 / SNARE complex.

Similar Datasets

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