A basic introduction to single particles cryo-electron microscopy

Vittoria Raimondi; Alessandro Grinzato; Vittoria Raimondi; Alessandro Grinzato

doi:10.3934/biophy.2022002

AIMS Biophysics

2022, Volume 9, Issue 1: 5-20. doi: 10.3934/biophy.2022002

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A basic introduction to single particles cryo-electron microscopy

Vittoria Raimondi ¹,
Alessandro Grinzato ^{2,3
,
,}

1.
Veneto Institute of Oncology IOV – IRCCS, via Gattamelata 64, 35128, Padova, Italy
2.
Department of Biomedical Sciences, University of Padova, Via Ugo Bassi 58b 35131 Padova, Italy
3.
Present address: ESRF, The European Synchrotron, 71 Avenue des Martyrs, 38043 Grenoble, France

Received: 31 October 2021 Revised: 25 November 2021 Accepted: 02 December 2021 Published: 30 December 2021

In the last years, cryogenic-electron microscopy (cryo-EM) underwent the most impressive improvement compared to other techniques used in structural biology, such as X-ray crystallography and NMR. Electron microscopy was invented nearly one century ago but, up to the beginning of the last decades, the 3D maps produced through this technique were poorly detailed, justifying the term “blobbology” to appeal to cryo-EM. Recently, thanks to a new generation of microscopes and detectors, more efficient algorithms, and easier access to computational power, single particles cryo-EM can routinely produce 3D structures at resolutions comparable to those obtained with X-ray crystallography. However, unlike X-ray crystallography, which needs crystallized proteins, cryo-EM exploits purified samples in solution, allowing the study of proteins and protein complexes that are hard or even impossible to crystallize. For these reasons, single-particle cryo-EM is often the first choice of structural biologists today. Nevertheless, before starting a cryo-EM experiment, many drawbacks and limitations must be considered. Moreover, in practice, the process between the purified sample and the final structure could be trickier than initially expected. Based on these observations, this review aims to offer an overview of the principal technical aspects and setups to be considered while planning and performing a cryo-EM experiment.
- Cryo-EM,
- electron microscope,
- sample preparation,
- data analysis
Citation: Vittoria Raimondi, Alessandro Grinzato. A basic introduction to single particles cryo-electron microscopy[J]. AIMS Biophysics, 2022, 9(1): 5-20. doi: 10.3934/biophy.2022002

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Abstract

In the last years, cryogenic-electron microscopy (cryo-EM) underwent the most impressive improvement compared to other techniques used in structural biology, such as X-ray crystallography and NMR. Electron microscopy was invented nearly one century ago but, up to the beginning of the last decades, the 3D maps produced through this technique were poorly detailed, justifying the term “blobbology” to appeal to cryo-EM. Recently, thanks to a new generation of microscopes and detectors, more efficient algorithms, and easier access to computational power, single particles cryo-EM can routinely produce 3D structures at resolutions comparable to those obtained with X-ray crystallography. However, unlike X-ray crystallography, which needs crystallized proteins, cryo-EM exploits purified samples in solution, allowing the study of proteins and protein complexes that are hard or even impossible to crystallize. For these reasons, single-particle cryo-EM is often the first choice of structural biologists today. Nevertheless, before starting a cryo-EM experiment, many drawbacks and limitations must be considered. Moreover, in practice, the process between the purified sample and the final structure could be trickier than initially expected. Based on these observations, this review aims to offer an overview of the principal technical aspects and setups to be considered while planning and performing a cryo-EM experiment.

Acknowledgments

We thanks Giuseppe Zanotti for reading and discussion of the draft.

^a regarding affiliation the writing work started at Università di Padova and ended at ESRF.

Conflict of interest

The authors declare no conflict of interest.

Author contributions:

AG concepted and designed the work, draft the article. VR edited and revised the manuscript.

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