Probing ligand binding of endothiapepsin by `temperature‐resolved' macromolecular crystallography

Continuous developments in cryogenic X‐ray crystallography have provided most of our knowledge of 3D protein structures, which has recently been further augmented by revolutionary advances in cryoEM. However, a single structural conformation identified at cryogenic temperatures may introduce a ficti...

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Published in:Acta crystallographica. Section D, Biological crystallography. Biological crystallography., 2022-08, Vol.78 (8), p.964-974
Main Authors: Huang, Chia-Ying, Aumonier, Sylvain, Engilberge, Sylvain, Eris, Deniz, Smith, Kate Mary Louise, Leonarski, Filip, Wojdyla, Justyna Aleksandra, Beale, John H., Buntschu, Dominik, Pauluhn, Anuschka, Sharpe, May Elizabeth, Metz, Alexander, Olieric, Vincent, Wang, Meitian
Format: Article
Language:English
Subjects:
Aspartic Acid Endopeptidases
Binding
conformational heterogeneity
Cryogenic cooling
Cryogenic effects
Cryogenic temperature
Crystallography
Crystallography, X-Ray
endothiapepsin
fragment binding
Ligands
Macromolecular Substances
Macromolecules
protein plasticity
Proteins
Proteins - chemistry
Research Papers
room temperature macromolecular crystallography
Temperature
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Summary:Continuous developments in cryogenic X‐ray crystallography have provided most of our knowledge of 3D protein structures, which has recently been further augmented by revolutionary advances in cryoEM. However, a single structural conformation identified at cryogenic temperatures may introduce a fictitious structure as a result of cryogenic cooling artefacts, limiting the overview of inherent protein physiological dynamics, which play a critical role in the biological functions of proteins. Here, a room‐temperature X‐ray crystallographic method using temperature as a trigger to record movie‐like structural snapshots has been developed. The method has been used to show how TL00150, a 175.15 Da fragment, undergoes binding‐mode changes in endothiapepsin. A surprising fragment‐binding discrepancy was observed between the cryo‐cooled and physiological temperature structures, and multiple binding poses and their interplay with DMSO were captured. The observations here open up new promising prospects for structure determination and interpretation at physiological temperatures with implications for structure‐based drug discovery. A room‐temperature X‐ray crystallographic method using temperature as a trigger to record movie‐like structural snapshots has been developed and applied to study ligand binding and protein plasticity.
ISSN:2059-7983
0907-4449
2059-7983
1399-0047
DOI:10.1107/S205979832200612X