Subtomogram analysis using the Volta phase plate

doi:10.1016/j.jsb.2016.05.009

. 2017 Feb;197(2):94-101.

doi: 10.1016/j.jsb.2016.05.009. Epub 2016 May 25.

Subtomogram analysis using the Volta phase plate

Maryam Khoshouei¹, Stefan Pfeffer¹, Wolfgang Baumeister¹, Friedrich Förster², Radostin Danev³

Affiliations

¹ Max-Planck Institute of Biochemistry, Department of Molecular Structural Biology, Am Klopferspitz 18, 82152 Martinsried, Germany.
² Max-Planck Institute of Biochemistry, Department of Molecular Structural Biology, Am Klopferspitz 18, 82152 Martinsried, Germany; Cryo-Electron Microscopy, Bijvoet Center for Biomolecular Research, Utrecht University, Padualaan 8, 3584 CH Utrecht, The Netherlands. Electronic address: foerster@biochem.mpg.de.
³ Max-Planck Institute of Biochemistry, Department of Molecular Structural Biology, Am Klopferspitz 18, 82152 Martinsried, Germany. Electronic address: danev@biochem.mpg.de.

PMID: 27235783
DOI: 10.1016/j.jsb.2016.05.009

Subtomogram analysis using the Volta phase plate

Maryam Khoshouei et al. J Struct Biol. 2017 Feb.

. 2017 Feb;197(2):94-101.

doi: 10.1016/j.jsb.2016.05.009. Epub 2016 May 25.

Authors

Maryam Khoshouei¹, Stefan Pfeffer¹, Wolfgang Baumeister¹, Friedrich Förster², Radostin Danev³

Affiliations

¹ Max-Planck Institute of Biochemistry, Department of Molecular Structural Biology, Am Klopferspitz 18, 82152 Martinsried, Germany.
² Max-Planck Institute of Biochemistry, Department of Molecular Structural Biology, Am Klopferspitz 18, 82152 Martinsried, Germany; Cryo-Electron Microscopy, Bijvoet Center for Biomolecular Research, Utrecht University, Padualaan 8, 3584 CH Utrecht, The Netherlands. Electronic address: foerster@biochem.mpg.de.
³ Max-Planck Institute of Biochemistry, Department of Molecular Structural Biology, Am Klopferspitz 18, 82152 Martinsried, Germany. Electronic address: danev@biochem.mpg.de.

PMID: 27235783
DOI: 10.1016/j.jsb.2016.05.009

Abstract

Cryo-electron tomography (CET) and subtomogram analysis allow studying the structures of macromolecular complexes in their natural context. The radiation sensitivity of vitrified biological specimens and the resulting low signal-to-noise ratio (SNR) in CET limit the amount of structural information that can be mined from tomographic data. The Volta phase plate (VPP) has emerged as an effective means to increase the SNR and hence contrast compared to 'conventional' defocus-based phase contrast transmission electron microscopy (CTEM). Here, we assess the performance of the VPP compared to CTEM in subtomogram analysis, using the mammalian 80S ribosome as a test case. Accurate focusing is the major factor for achieving high resolution with the VPP, as highlighted by a comparison of slightly different focusing strategies. From only 1400 subtomograms, the VPP yields a subtomogram average of the mammalian 80S ribosome at 9.6Å resolution without laborious contrast transfer function (CTF) correction. The subtomogram averages obtained using CTEM approaches are comparable, but suffer from lower signal transfer in certain frequency bands due to the oscillations of the CTF. Our study demonstrates that the VPP is a valuable tool for subtomogram analysis, because it enables improved performance and efficiency in terms of structure localization and number of subtomograms required for a given resolution.

Keywords: Cryo-electron tomography; Ribosome; Subtomogram analysis; Volta phase plate.

PubMed Disclaimer

Cited by

Beam image-shift accelerated data acquisition for near-atomic resolution single-particle cryo-electron tomography.
Bouvette J, Liu HF, Du X, Zhou Y, Sikkema AP, da Fonseca Rezende E Mello J, Klemm BP, Huang R, Schaaper RM, Borgnia MJ, Bartesaghi A. Bouvette J, et al. Nat Commun. 2021 Mar 30;12(1):1957. doi: 10.1038/s41467-021-22251-8. Nat Commun. 2021. PMID: 33785757 Free PMC article.
High-resolution structure determination using high-throughput electron cryo-tomography.
Liu HF, Zhou Y, Bartesaghi A. Liu HF, et al. Acta Crystallogr D Struct Biol. 2022 Jul 1;78(Pt 7):817-824. doi: 10.1107/S2059798322005010. Epub 2022 Jun 7. Acta Crystallogr D Struct Biol. 2022. PMID: 35775981 Free PMC article. Review.
Entropy-regularized deconvolution of cellular cryotransmission electron tomograms.
Croxford M, Elbaum M, Arigovindan M, Kam Z, Agard D, Villa E, Sedat J. Croxford M, et al. Proc Natl Acad Sci U S A. 2021 Dec 14;118(50):e2108738118. doi: 10.1073/pnas.2108738118. Proc Natl Acad Sci U S A. 2021. PMID: 34876518 Free PMC article.
Multi-particle cryo-EM refinement with M visualizes ribosome-antibiotic complex at 3.5 Å in cells.
Tegunov D, Xue L, Dienemann C, Cramer P, Mahamid J. Tegunov D, et al. Nat Methods. 2021 Feb;18(2):186-193. doi: 10.1038/s41592-020-01054-7. Epub 2021 Feb 4. Nat Methods. 2021. PMID: 33542511 Free PMC article.
Accurate size-based protein localization from cryo-ET tomograms.
Jin W, Zhou Y, Bartesaghi A. Jin W, et al. J Struct Biol X. 2024 Jun 26;10:100104. doi: 10.1016/j.yjsbx.2024.100104. eCollection 2024 Dec. J Struct Biol X. 2024. PMID: 39044770 Free PMC article.

See all "Cited by" articles

Publication types

Actions

MeSH terms

Actions
Actions
Actions
Actions
Actions
Actions
Actions
Actions
Actions
Actions

LinkOut - more resources

Full Text Sources
- Elsevier Science
Other Literature Sources
- scite Smart Citations

Save citation to file

Email citation

Add to Collections

Add to My Bibliography

Your saved search

Create a file for external citation management software

Your RSS Feed

Subtomogram analysis using the Volta phase plate

Affiliations

Subtomogram analysis using the Volta phase plate

Authors

Affiliations

Abstract

Similar articles

Cited by

Publication types

MeSH terms

LinkOut - more resources

Full Text Sources

Other Literature Sources