Titan Krios; Structural Biology; Cryo-electron microscopy; Macromolecular structure; Cell biology; Single particle analysis; Electron cryo-tomography
Bloch Joël S., Pesciullesi Giorgio, Boilevin Jérémy, Nosol Kamil, Irobalieva Rossitza N., Darbre Tamis, Aebi Markus, Kossiakoff Anthony A., Reymond Jean-Louis, Locher Kaspar P. (2020), Structure and mechanism of the ER-based glucosyltransferase ALG6, in Nature
, 579(7799), 443-447.
Zachs Tobias, Schertel Andreas, Medeiros João, Weiss Gregor L, Hugener Jannik, Matos Joao, Pilhofer Martin (2020), Fully automated, sequential focused ion beam milling for cryo-electron tomography, in eLife
, 9, 0-0.
Olsen Jeppe A., Alam Amer, Kowal Julia, Stieger Bruno, Locher Kaspar P. (2020), Structure of the human lipid exporter ABCB4 in a lipid environment, in Nature Structural & Molecular Biology
, 27(1), 62-70.
Ramírez Ana S., Kowal Julia, Locher Kaspar P. (2019), Cryo–electron microscopy structures of human oligosaccharyltransferase complexes OST-A and OST-B, in Science
, 366(6471), 1372-1375.
Eisenstein Fabian, Danev Radostin, Pilhofer Martin (2019), Improved applicability and robustness of fast cryo-electron tomography data acquisition, in Journal of Structural Biology
, 208(2), 107-114.
Qi Chao, Di Minin Giulio, Vercellino Irene, Wutz Anton, Korkhov Volodymyr M. (2019), Structural basis of sterol recognition by human hedgehog receptor PTCH1, in Science Advances
, 5(9), eaaw6490-eaaw6490.
Visualizing biological macromolecules and cellular structures at high resolution is of key importance for understanding their functions in biology. Recently, a revolution has propelled cryo-electron microscopy (cryo-EM) to the forefront of structural biology by increasing the resolution with which cellular structures can be studied. As a consequence, single particle cryo-EM and electron cryo-tomography have enormously gained in importance and are widely used to study the structures of macromolecular complexes at near-atomic resolution. Researchers worldwide are applying this method to investigate macromolecules or complexes that are not amenable to X-ray crystallography or NMR. This also applies to researchers at ETH and in Switzerland, many of whom now critically depend on cryo-EM for their explorations of fundamental questions of cellular life.Housed within the ScopeM platform, ETH operates a high-end Titan Krios cryo-electron microscope that is currently upgraded to allow both single particle and tomography experiments and allow visualization of smaller particles (<200kDa) than previously possible. However, the capacity of this instrument is limited, rendering efficient sample analysis and optimization critical. With the present proposal, the applicants wish to drastically increase the capacity for cryo-EM studies by acquiring a Talos Arctica instrument. The goal for this 200kV cryo-EM instrument is to facilitate the following: (i) Efficient analysis of the homogeneity of single particle samples by fully automated, unsupervised data acquisition; (ii) Single-particle visualization not only of large complexes, but also smaller particles, which can be achieved with a Gatan K2XP direct detector; (iii) Cryo-tomography studies must be possible, for which the instrument will be equipped with a Gatan Imaging Filter (GIF); (iv) Finally, the instrument should feature an autoloader whose autogrids are compatible with that of the Titan Krios, allowing direct sample transfer for high-value samples. The Talos Arctica microscope is the only instrument currently available that provides the required features.The Talos Arctica cryo-EM instrument will be operated within the ETH ScopeM platform, which provides access to all research groups requiring cryo-EM access. The individual projects of the applicants outline the many research topics that will benefit from the new instrument. These include, on the one hand, single particle cryo-EM studies of large macromolecular protein-protein or protein-nucleic acide complexes mediating key physiological processes, but also of much smaller membrane proteins that facilitate critical transport or receptor functions. Single particle analysis will also allow engineered and naturally occurring protein containers to be studied, as well as providing structural data to interpret proteome analyses. In addition, the Talos Arctica will allow advanced cryo-tomographic projects to be pursued. These deal, among others, with nucleopores, bacterial cell interactions, and intracellular starch crystallization. In addition to the applicants, there are additional groups at ETH whose research requires access to cryo-EM instruments.The acquisition of a Talos Arctica cryo-EM intrument will alleviate a critical bottleneck that currently exists for many highly competitive projects. The output will critically contribute to fundamental research in life sciences, but also stimulate hybrid approaches in structural biology, and greatly impact our understanding of disease mechanisms.