Journal
JOURNAL OF STRUCTURAL BIOLOGY
Volume 208, Issue 2, Pages 107-114Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.jsb.2019.08.006
Keywords
Cryo-electron tomography; Electron cryotomography; Cryo-EM; Tilt series; Data acquisition; Throughput; K2; K3; Calibration
Funding
- ETH Zurich
- NOMIS foundation
- Swiss National Science Foundation [164092, 179255, 170808]
- European Research Council [679209]
- Japan Society for the Promotion of Science (JSPS) KAKENHI [18H06043]
- Japan Science and Technology Agency (JST) PRESTO [18069571]
- Japan Society for the Promotion of Science (JSPS)
- Grants-in-Aid for Scientific Research [18H06043] Funding Source: KAKEN
- European Research Council (ERC) [679209] Funding Source: European Research Council (ERC)
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The power of cryo-electron tomography (cryoET) lies in its capability to characterize macromolecules in their cellular context. Structure determination by cryoET, however, is time-consuming compared to single particle approaches. A recent study reported significant acceleration of data acquisition by a fast-incremental single-exposure (FISE) tilt series scheme. Here we improved the method and evaluated its efficiency and performance. We show that (1) FISE combined with the latest generation of direct electron detectors speeds up collection considerably, (2) previous generation (pre-2017) double-tilt axis Titan Krios holders are also suitable for FISE data acquisition, (3) x, y and z-specimen shifts can be compensated for, and (4) FISE tilt series data can generate averages of sub-nanometer resolution. These advances will allow for a widespread adoption of cryoET for high-throughput in situ studies and high-resolution structure determination across different biological research disciplines.
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