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Protein-to-structure pipeline for ambient-temperature in situ crystallography at VMXi

DOI: 10.1107/S2052252523003810 DOI Help

Authors: Halina Mikolajek (Diamond Light Source) , Juan Sanchez-Weatherby (Diamond Light Source) , James Sandy (Diamond Light Source) , Richard J. Gildea (Diamond Light Source) , Ivan Campeotto (Nottingham Trent University; University of Nottingham) , Harish Cheruvara (Research Complex at Harwell (RCaH); Diamond Light Source) , John D. Clarke (Diamond Light Source; University of Oxford; Pirbright Institute; The Rosalind Franklin Institute) , Toshana Foster (University of Nottingham) , Sotaro Fujii (Diamond Light Source; Hiroshima University) , Ian T. Paulsen (Macquarie University) , Bhumika S. Shah (Macquarie University) , Michael A. Hough (Diamond Light Source)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Iucrj , VOL 10

State: Published (Approved)
Published: July 2023

Open Access Open Access

Abstract: The utility of X-ray crystal structures determined under ambient-temperature conditions is becoming increasingly recognized. Such experiments can allow protein dynamics to be characterized and are particularly well suited to challenging protein targets that may form fragile crystals that are difficult to cryo-cool. Room-temperature data collection also enables time-resolved experiments. In contrast to the high-throughput highly automated pipelines for determination of structures at cryogenic temperatures widely available at synchrotron beamlines, room-temperature methodology is less mature. Here, the current status of the fully automated ambient-temperature beamline VMXi at Diamond Light Source is described, and a highly efficient pipeline from protein sample to final multi-crystal data analysis and structure determination is shown. The capability of the pipeline is illustrated using a range of user case studies representing different challenges, and from high and lower symmetry space groups and varied crystal sizes. It is also demonstrated that very rapid structure determination from crystals in situ within crystallization plates is now routine with minimal user intervention.

Journal Keywords: room temperature; in situ; multi-crystal; crystallization pipelines; automated data collection; structural biology; radiation damage; X-ray crystallography; VMXi

Subject Areas: Biology and Bio-materials

Instruments: VMXi-Versatile Macromolecular Crystallography in situ

Added On: 22/05/2023 10:49


Discipline Tags:

Structural biology Life Sciences & Biotech

Technical Tags:

Diffraction Macromolecular Crystallography (MX)