Below 3 Å structure of apoferritin using a multipurpose TEM with a side entry cryoholder

Abstract Recently, the structural analysis of protein complexes by cryo-electron microscopy (cryo-EM) single particle analysis (SPA) has had great impact as a biophysical method. Many results of cryo-EM SPA are based on data acquired on state-of-the-art cryo-electron microscopes customized for SPA....

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Autores principales:	Yoko Kayama, Raymond N. Burton-Smith, Chihong Song, Naoya Terahara, Takayuki Kato, Kazuyoshi Murata
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/824cd0b7e1fc490baf90f488d0657eb1
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spelling	oai:doaj.org-article:824cd0b7e1fc490baf90f488d0657eb12021-12-02T14:26:12ZBelow 3 Å structure of apoferritin using a multipurpose TEM with a side entry cryoholder10.1038/s41598-021-87183-12045-2322https://doaj.org/article/824cd0b7e1fc490baf90f488d0657eb12021-04-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-87183-1https://doaj.org/toc/2045-2322Abstract Recently, the structural analysis of protein complexes by cryo-electron microscopy (cryo-EM) single particle analysis (SPA) has had great impact as a biophysical method. Many results of cryo-EM SPA are based on data acquired on state-of-the-art cryo-electron microscopes customized for SPA. These are currently only available in limited locations around the world, where securing machine time is highly competitive. One potential solution for this time-competitive situation is to reuse existing multi-purpose equipment, although this comes with performance limitations. Here, a multi-purpose TEM with a side entry cryo-holder was used to evaluate the potential of high-resolution SPA, resulting in a 3 Å resolution map of apoferritin with local resolution extending to 2.6 Å. This map clearly showed two positions of an aromatic side chain. Further, examination of optimal imaging conditions depending on two different multi-purpose electron microscope and camera combinations was carried out, demonstrating that higher magnifications are not always necessary or desirable. Since automation is effectively a requirement for large-scale data collection, and augmenting the multi-purpose equipment is possible, we expanded testing by acquiring data with SerialEM using a β-galactosidase test sample. This study demonstrates the possibilities of more widely available and established electron microscopes, and their applications for cryo-EM SPA.Yoko KayamaRaymond N. Burton-SmithChihong SongNaoya TeraharaTakayuki KatoKazuyoshi MurataNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-13 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Yoko Kayama Raymond N. Burton-Smith Chihong Song Naoya Terahara Takayuki Kato Kazuyoshi Murata Below 3 Å structure of apoferritin using a multipurpose TEM with a side entry cryoholder
description	Abstract Recently, the structural analysis of protein complexes by cryo-electron microscopy (cryo-EM) single particle analysis (SPA) has had great impact as a biophysical method. Many results of cryo-EM SPA are based on data acquired on state-of-the-art cryo-electron microscopes customized for SPA. These are currently only available in limited locations around the world, where securing machine time is highly competitive. One potential solution for this time-competitive situation is to reuse existing multi-purpose equipment, although this comes with performance limitations. Here, a multi-purpose TEM with a side entry cryo-holder was used to evaluate the potential of high-resolution SPA, resulting in a 3 Å resolution map of apoferritin with local resolution extending to 2.6 Å. This map clearly showed two positions of an aromatic side chain. Further, examination of optimal imaging conditions depending on two different multi-purpose electron microscope and camera combinations was carried out, demonstrating that higher magnifications are not always necessary or desirable. Since automation is effectively a requirement for large-scale data collection, and augmenting the multi-purpose equipment is possible, we expanded testing by acquiring data with SerialEM using a β-galactosidase test sample. This study demonstrates the possibilities of more widely available and established electron microscopes, and their applications for cryo-EM SPA.
format	article
author	Yoko Kayama Raymond N. Burton-Smith Chihong Song Naoya Terahara Takayuki Kato Kazuyoshi Murata
author_facet	Yoko Kayama Raymond N. Burton-Smith Chihong Song Naoya Terahara Takayuki Kato Kazuyoshi Murata
author_sort	Yoko Kayama
title	Below 3 Å structure of apoferritin using a multipurpose TEM with a side entry cryoholder
title_short	Below 3 Å structure of apoferritin using a multipurpose TEM with a side entry cryoholder
title_full	Below 3 Å structure of apoferritin using a multipurpose TEM with a side entry cryoholder
title_fullStr	Below 3 Å structure of apoferritin using a multipurpose TEM with a side entry cryoholder
title_full_unstemmed	Below 3 Å structure of apoferritin using a multipurpose TEM with a side entry cryoholder
title_sort	below 3 å structure of apoferritin using a multipurpose tem with a side entry cryoholder
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/824cd0b7e1fc490baf90f488d0657eb1
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Below 3 Å structure of apoferritin using a multipurpose TEM with a side entry cryoholder

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