Fiche publication
Date publication
octobre 2022
Journal
Journal of structural biology
Auteurs
Membres identifiés du Cancéropôle Est :
Dr KLAHOLZ Bruno
Tous les auteurs :
Fréchin L, Holvec S, von Loeffelholz O, Hazemann I, Klaholz BP
Lien Pubmed
Résumé
Recent technological advances in cryo electron microscopy (cryo-EM) have led to new opportunities in the structural biology field. Here we benchmark the performance of two 300 kV latest-generation cryo electron microscopes, Titan Krios G4 from Thermofisher Scientific and CRYO ARM 300 from Jeol, with regards to achieving high resolution single particle reconstructions on a real case sample. We compare potentially limiting factors such as drift rates, astigmatism & coma aberrations and performance during image processing and show that both microscopes, while comprising rather different technical setups & parameter settings and equipped with different types of energy filters & cameras, achieve a resolution of around 2 Å on the human ribosome, a non-symmetric object which constitutes a key drug target. Astigmatism correction, CTF refinement and correction of higher order aberrations through refinement in separate optics groups helped to account for astigmatism/coma caused by beam tilting during multi-spot and multi-hole acquisition in neighbouring holes without stage movement. The obtained maps resolve Mg ions, water molecules, inhibitors and side-chains including chemical modifications. The fact that both instruments can resolve such features will greatly facilitate understanding molecular mechanisms and helps in cryo-EM structure based drug design. The methods and analysis tools used here will be useful also to characterize existing instruments and optimize data acquisition settings and are applicable broadly to other drug targets in structural biology.
Référence
J Struct Biol. 2022 10 11;:107905
