8AWX

Millisecond cryo-trapping by the spitrobot crystal plunger, Xylose Isomerase with Glucose at 1s

Summary for 8AWX

• Entry DOI: 10.2210/pdb8awx/pdb
• Descriptor: Xylose isomerase, MAGNESIUM ION, beta-D-glucopyranose, ... (5 entities in total)
• Functional Keywords: xylose isomerase, glucose isomerase, humidity, space group change, unit cell change, time-resolved crystallography, isomerase
• Biological source: Streptomyces rubiginosus
• Total number of polymer chains: 1
• Total formula weight: 43542.70

Authors

Mehrabi, P.,Sung, S.,von Stetten, D.,Prester, A.,Hatton, C.E.,Kleine-Doepke, S.,Berkes, A.,Gore, G.,Leimkohl, J.P.,Schikora, H.,Kollewe, M.,Rohde, H.,Wilmanns, M.,Tellkamp, F.,Schulz, E.C. (deposition date: 2022-08-30, release date: 2023-05-24, Last modification date: 2024-02-07)

Primary citation

Mehrabi, P.,Sung, S.,von Stetten, D.,Prester, A.,Hatton, C.E.,Kleine-Dopke, S.,Berkes, A.,Gore, G.,Leimkohl, J.P.,Schikora, H.,Kollewe, M.,Rohde, H.,Wilmanns, M.,Tellkamp, F.,Schulz, E.C.
Millisecond cryo-trapping by the spitrobot crystal plunger simplifies time-resolved crystallography.
Nat Commun, 14:2365-2365, 2023

PubMed 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.

PubMed: 37185266
DOI: 10.1038/s41467-023-37834-w

Experimental method

X-RAY DIFFRACTION (1.96 Å)