Millisecond cryo-trapping by the spitrobot crystal plunger simplifies 
time-resolved crystallography

Mehrabi, P.; Sung, S.; von Stetten, D.; Prester, A.; Hatton, C. E.; Kleine-Döpke, S.; Berkes, A.; Gore, G.; Leimkohl, J.-P.; Schikora, H.; Kollewe, M.; Rohde, H.; Wilmanns, M.; Tellkamp, F.; Schulz, E.-C.

doi:10.1038/s41467-023-37834-w

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Millisecond cryo-trapping by the spitrobot crystal plunger simplifies time-resolved crystallography

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Mehrabi, P.
Institute for Nanostructure and Solid State Physics, Universität Hamburg;
Miller Group, Atomically Resolved Dynamics Department, Max Planck Institute for the Structure and Dynamics of Matter, Max Planck Society;

/persons/resource/persons241300

Leimkohl, J.-P.
Machine Physics, Scientific Service Units, Max Planck Institute for the Structure and Dynamics of Matter, Max Planck Society;

/persons/resource/persons241303

Schikora, H.
Machine Physics, Scientific Service Units, Max Planck Institute for the Structure and Dynamics of Matter, Max Planck Society;

/persons/resource/persons281287

Kollewe, M.
Machine Physics, Scientific Service Units, Max Planck Institute for the Structure and Dynamics of Matter, Max Planck Society;

/persons/resource/persons182596

Tellkamp, F.
Machine Physics, Scientific Service Units, Max Planck Institute for the Structure and Dynamics of Matter, Max Planck Society;

/persons/resource/persons202768

Schulz, E.-C.
Institute for Nanostructure and Solid State Physics, Universität Hamburg;
Miller Group, Atomically Resolved Dynamics Department, Max Planck Institute for the Structure and Dynamics of Matter, Max Planck Society;
University Medical Center Hamburg-Eppendorf (UKE);

External Resource

https://doi.org/10.1101/2022.09.20.508674
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https://doi.org/10.1038/s41467-023-37834-w
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Citation

Mehrabi, P., Sung, S., von Stetten, D., Prester, A., Hatton, C. E., Kleine-Döpke, S., et al. (2023). Millisecond cryo-trapping by the spitrobot crystal plunger simplifies time-resolved crystallography. Nature Communications, 14(1): 2365. doi:10.1038/s41467-023-37834-w.

Cite as: https://hdl.handle.net/21.11116/0000-000B-3265-B

Abstract

We introduce the spitrobot, a protein crystal plunger, enabling reaction quenching via cryo-trapping with a time-resolution in the millisecond range. Protein crystals are mounted on canonical micromeshes on an electropneumatic piston, where the crystals are kept in a humidity and temperature-controlled environment, then reactions are initiated via the liquid application method (LAMA) and plunging into liquid nitrogen is initiated after an electronically set delay time to cryo-trap intermediate states. High-magnification images are automatically recorded before and after droplet deposition, prior to plunging. The SPINE-standard sample holder is directly plunged into a storage puck, enabling compatibility with high-throughput infrastructure. Here we demonstrate binding of glucose and 2,3-butanediol in microcrystals of xylose isomerase, and of avibactam and ampicillin in microcrystals of the extended spectrum beta-lactamase CTX-M-14. We also trap reaction intermediates and conformational changes in macroscopic crystals of tryptophan synthase to demonstrate that the spitrobot enables insight into catalytic events.