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Acquisition of a Talos Arctica transmission electron microscope for single particle analysis and cryo-tomography

English title Acquisition of a Talos Arctica transmission electron microscope for single particle analysis and cryo-tomography
Applicant Locher Kaspar
Number 170808
Funding scheme R'EQUIP
Research institution Institut für Molekularbiologie und Biophysik Deptartement für Biologie ETH Zurich
Institution of higher education ETH Zurich - ETHZ
Main discipline Biophysics
Start/End 01.05.2017 - 30.04.2020
Approved amount 950'000.00
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All Disciplines (2)

Discipline
Biophysics
Biochemistry

Keywords (7)

Titan Krios; Structural Biology; Cryo-electron microscopy; Macromolecular structure; Cell biology; Single particle analysis; Electron cryo-tomography

Lay Summary (German)

Lead
Mit der Cryo-Elektronenmikroskopie können biologische Makromoleküle sowie Zellkomponenten mit hoher Auflösung dargestellt werden. Diese Technik revolutioniert gegenwärtig die molekulare Biologie. Mit der Anschaffung eines Cryo-Elektronenmikroskop Titan Krios erhalten wir Anschluss an diese Spitzentechnologie.
Lay summary

Cryo-Elektronenmikroskopie (cryo-EM) hat sich zu einer der wichtigsten Methoden entwickelt, zelluläre Strukturen und biologische Makromoleküle zu erforschen. Die Technik, biologische Proben im gefrorenen Zustand mit einem Transmissions-Elektronenmikroskop (TEM) zu untersuchen, wurde zwar schon vor mehreren Jahrzenten entwickelt und hat die life sciences seit da nachhaltig beeinflusst. Die Entwicklung einer neuen Generation von Detektoren hat vor kurzem eine Revolution eingeleitet, die cryo-EM zur gegenwärtig erfolgreichsten Methode transformiert, die Komponenten lebender Zellen mit hoher Auflösung darzustellen. Cryo-EM ist damit im Begriff, Röntgenkristallographie als bislang erfolgreichste Methode zur de novo Strukturbestimmung biologischer Makromoleküle abzulösen.

Forscher der ETH Zürich werden mit dem Titan Krios Mikroskop die Mechanismen vieler biologisch und biomedizinisch relevanter Prozesse untersuchen können. Im Gegensatz zur  Röntgenkristallographie können mit dem Spitzenmikroskop "Titan Krios" die Strukturen von Einzelmolekülen (single particle analysis) sowie ganze Zellen mittel Tomographie dargestellt werden, und zwar mit einer Auflösung, die eine atomare Interpretation zulässt, eine Domäne, die bislang nur mittels Röntgenkristallographie erreicht werden konnte. Als Folge dieser Entwicklung bemühen sich weltweit alle Spitzenuniversitäten mit enormem Eifer und Aufwand, ihre Cryo-EM Kapazitäten auszubauen, um der akademische Forschung den Zugang zu dieser Schlüsseltechnologie zu ermöglichen. Aber auch die pharmazeutische Industrie investiert in diesen Bereich, da die Technik auch der Medikamentenentwicklung entscheidende neue Impulse geben wird. Das Cryo-EM wird sich ebenfalls zu einem wichtigen Pfeiler der "personalized medicine" entwickeln, da sie effiziente Strukturuntersuchungen mit kleinsten Mengen Material ermöglicht.

Direct link to Lay Summary Last update: 24.05.2017

Responsible applicant and co-applicants

Publications

Publication
Structure and mechanism of the ER-based glucosyltransferase ALG6
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.
Fully automated, sequential focused ion beam milling for cryo-electron tomography
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.
Structure of the human lipid exporter ABCB4 in a lipid environment
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.
Cryo–electron microscopy structures of human oligosaccharyltransferase complexes OST-A and OST-B
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.
Improved applicability and robustness of fast cryo-electron tomography data acquisition
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.
Structural basis of sterol recognition by human hedgehog receptor PTCH1
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.

Associated projects

Number Title Start Funding scheme
141735 NCCR RNA & disease: Understanding the role of RNA biology in disease mechanisms (phase I) 01.05.2014 National Centres of Competence in Research (NCCRs)
147632 Transglyco: Chemistry, Enzymology and Physiology of Oligosaccharyltransferase 01.11.2013 Sinergia
163314 The pupylation-dependent and -independent proteasome pathways of mycobacteria 01.11.2015 Project funding
156304 Functional significance of the dynamics of receptor binding and the alternative folding possibilities of pilus subunits in urinary tract infections caused by pathogenic Escherichia coli strains 01.10.2014 Project funding
144035 In vivo and in vitro structural analysis of flagellar/ciliary motors and regulators 01.07.2013 Project funding
166617 3D cryo-EM analysis of the axoneme and the transition zone from cilia 01.03.2017 Project funding
163503 Designing starch - harnessing carbohydrate polymer synthesis in plants 01.10.2015 ERA-CAPS
125762 NCCR TransCure: From transport physiology to identification of therapeutic targets (phase I) 01.11.2010 National Centres of Competence in Research (NCCRs)
150665 Structural molecular biology of membrane proteins in signal transduction and cholesterol recognition 01.04.2014 SNSF Professorships
152878 Structure and Mechanism of Bacterial Contractile Assemblies 01.12.2014 Project funding
189111 Structural and mechanistic studies of liver ABC transporters 01.10.2019 Project funding
200294 Systematic cryo-EM and proteomic analysis of protein complexes related to primary ciliary dyskinesia 01.11.2021 India
192644 Structural study on calcium regulation of motile cilia by cryo-EM 01.04.2020 Project funding
149921 NMR structure determination of protein-RNA complexes involved in pre-mRNA splicing and translation regulation 01.10.2013 Project funding
153144 Understanding the Mechanism of Starch Biosynthesis in Plants 01.04.2014 Project funding
166672 Structural and mechanistic studies of components of bacterial protein N-glycosylation pathway and of vitamin B12 transport 01.04.2016 Project funding
159731 Structure and Function of the Nuclear Pore Complex 01.04.2015 Project funding
163478 Bonus of Excellence - Structural studies of complexes involved in ribosome assembly and translation initiation in yeast 01.11.2015 Project funding
164092 Upgrade of a Titan Krios electron cryo-microscope for single particle analysis and tomography 01.12.2015 R'EQUIP

Abstract

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.
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