Abstract
Serial crystallography has enabled the study of complex biological questions through the determination of biomolecular structures at room temperature using low X-ray doses. Furthermore, it has enabled the study of protein dynamics by the capture of atomically resolved and time-resolved molecular movies. However, the study of many biologically relevant targets is still severely hindered by high sample consumption and lengthy data-collection times. By combining serial synchrotron crystallography (SSX) with 3D printing, a new experimental platform has been created that tackles these challenges. An affordable 3D-printed, X-ray-compatible microfluidic device (3D-MiXD) is reported that allows data to be collected from protein microcrystals in a 3D flow with very high hit and indexing rates, while keeping the sample consumption low. The miniaturized 3D-MiXD can be rapidly installed into virtually any synchrotron beamline with only minimal adjustments. This efficient collection scheme in combination with its mixing geometry paves the way for recording molecular movies at synchrotrons by mixing-triggered millisecond time-resolved SSX.
Highlights
Serial synchrotron crystallography (SSX) is a data-collection approach in which diffraction data are collected from lowmillisecond X-ray exposures of protein microcrystals
The 3D-MiXD designed, fabricated and used in this work is shown in Fig. 1(a)
The most important aspect of the device design was the introduction of a 3D flowfocusing geometry, where the sample is focused in the center of the channel by buffer or water
Summary
Serial synchrotron crystallography (SSX) is a data-collection approach in which diffraction data are collected from lowmillisecond X-ray exposures of protein microcrystals. One of the main advantages of this technique is the low X-ray dose that is accumulated by the crystals during data collection, as fresh crystalline material is available for each new exposure (Ebrahim et al, 2019; Owen et al, 2017). Multi-crystal experiments have been carried out in the past for very radiation-sensitive samples such as viruses (Abrescia et al, 2004; Ji et al, 2009), the high brilliance of third- and fourth-generation synchrotrons and X-ray free-electron laser (XFEL) sources has propelled the recent rapid development of serial crystallography. Data-acquisition Crystal size Indexing 5000 indexed rate (Hz).
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