Fixed-Target Serial Synchrotron Crystallography Using Nylon Mesh and Enclosed Film-Based Sample Holder

Yunje Cho,Hyeongju Choi,Ki Hyun Nam,Suk-Youl Park,Cheolsoo Eo

doi:10.3390/cryst10090803

Yunje Cho, Hyeongju Choi + Show 3 more

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https://doi.org/10.3390/cryst10090803

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Journal: Crystals	Publication Date: Sep 10, 2020
Citations: 24	License type: CC BY 4.0

Affiliation: Pohang University of Science and Technology

Abstract

Serial crystallography (SX) technique using synchrotron X-ray allows the visualization of room-temperature crystal structures with low-dose data collection as well as time-resolved molecular dynamics. In an SX experiment, delivery of numerous crystals for X-ray interaction, in a serial manner, is very important. Fixed-target scanning approach has the advantage of dramatically minimizing sample consumption as well as any physical damage to crystal sample, compared to other sample delivery methods. Here, we introduce the simple approach of fixed-target serial synchrotron crystallography (FT-SSX) using nylon mesh and enclosed film (NAM)-based sample holder. The NAM-based sample holder consisted of X-ray-transparent nylon-mesh and polyimide film, attached to a magnetic base. This sample holder was mounted to a goniometer head on macromolecular crystallography beamline, and translated along vertical and horizontal directions for raster scanning by the goniometer. Diffraction data were collected in two raster scanning approaches: (i) 100 ms X-ray exposure and 0.011° oscillation at each scan point and (ii) 500 ms X-ray exposure and 0.222° oscillation at each scan point. Using this approach, we determined the room-temperature crystal structures of lysozyme and glucose isomerase at 1.5–2.0 Å resolution. The sample holder produced negligible X-ray background scattering for data processing. Therefore, the new approach provided an opportunity to perform FT-SSX with high accessibility using macromolecular crystallography beamlines at synchrotron without any special equipment.

Highlights

Traditional X-ray crystallography is a powerful tool for understanding the biological functions of macromolecules at atomic resolution [1]
Be overcome by serial crystallography (SX) techniques, using X-ray free-electron laser (XFEL) or synchrotron X-ray, which allow the observation of crystal structure at room temperature, with minimal radiation damage [9,10]
This sample holder was mounted on a translation stage in the FT-SFX chamber to allow scanning of a fixed target and collect diffraction data [36]

Summary

Introduction

Traditional X-ray crystallography is a powerful tool for understanding the biological functions of macromolecules at atomic resolution [1]. The structural information could often be biologically irrelevant [7,8] Such experimental limitation can, be overcome by serial crystallography (SX) techniques, using X-ray free-electron laser (XFEL) or synchrotron X-ray, which allow the observation of crystal structure at room temperature, with minimal radiation damage [9,10]. Be overcome by serial crystallography (SX) techniques, using X-ray free-electron laser (XFEL) or synchrotron X-ray, which allow the observation of crystal structure at room temperature, with minimal radiation damage [9,10] Such techniques allow the visualization of time-resolved molecular dynamics through a pump-probe experiment using an optical laser or liquid application [10,11,12,13,14,15]

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Discussion

Conclusion