GraFix: Sample preparation for single-particle electron cryomicroscopy

Berthold Kastner; Niels Fischer; Monika Mariola Golas; Bjoern Sander; Prakash Dube; Daniel Boehringer; Klaus Hartmuth; Jochen Deckert; Florian Hauer; Elmar Wolf; Hannes Uchtenhagen; Henning Urlaub; Franz Herzog; Jan Michael Peters; Dietmar Poerschke; Reinhard Lührmann; Holger Stark

doi:10.1038/nmeth1139

GraFix: Sample preparation for single-particle electron cryomicroscopy

Berthold Kastner, Niels Fischer, Monika Mariola Golas, Bjoern Sander, Prakash Dube, Daniel Boehringer, Klaus Hartmuth, Jochen Deckert, Florian Hauer, Elmar Wolf, Hannes Uchtenhagen, Henning Urlaub, Franz Herzog, Jan Michael Peters, Dietmar Poerschke, Reinhard Lührmann, Holger Stark

Publikation: Beitrag in Fachzeitschrift › Artikel › Begutachtung

Abstract

We developed a method, named GraFix, that considerably improves sample quality for structure determination by single-particle electron cryomicroscopy (cryo-EM). GraFix uses a glycerol gradient centrifugation step in which the complexes are centrifuged into an increasing concentration of a chemical fixation reagent to prevent aggregation and to stabilize individual macromolecules. The method can be used to prepare samples for negative-stain, cryo-negative-stain and, particularly, unstained cryo-EM.

Originalsprache	Englisch
Seiten (von - bis)	53-55
Seitenumfang	3
Fachzeitschrift	Nature Methods
Jahrgang	5
Ausgabenummer	1
DOIs	https://doi.org/10.1038/nmeth1139
Publikationsstatus	Veröffentlicht - Jän. 2008
Extern publiziert	Ja

Forschungsfelder

Chemical Crosslinking
Structural Biology

IMC Forschungsschwerpunkte

Medical biotechnology

ÖFOS 2012 - Österreichischen Systematik der Wissenschaftszweige

106037 Proteomik
106041 Strukturbiologie
106044 Systembiologie

UN SDGs

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10.1038/nmeth1139

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title = "GraFix: Sample preparation for single-particle electron cryomicroscopy",

abstract = "We developed a method, named GraFix, that considerably improves sample quality for structure determination by single-particle electron cryomicroscopy (cryo-EM). GraFix uses a glycerol gradient centrifugation step in which the complexes are centrifuged into an increasing concentration of a chemical fixation reagent to prevent aggregation and to stabilize individual macromolecules. The method can be used to prepare samples for negative-stain, cryo-negative-stain and, particularly, unstained cryo-EM.",

author = "Berthold Kastner and Niels Fischer and Golas, {Monika Mariola} and Bjoern Sander and Prakash Dube and Daniel Boehringer and Klaus Hartmuth and Jochen Deckert and Florian Hauer and Elmar Wolf and Hannes Uchtenhagen and Henning Urlaub and Franz Herzog and Peters, {Jan Michael} and Dietmar Poerschke and Reinhard L{\"u}hrmann and Holger Stark",

note = "Funding Information: We thank H. Kohansal, T. Conrad and W. Jahn for expert technical assistance, U. G{\"o}ringer and M. Brecht for preparing the RNA editing complex, M. Rodnina and K. Gromadski for preparing the 70S ribosome complex, and C. Will for preparing the 17S U2 snRNP. The work was supported by grants from the Federal Ministry of Education and Research, Germany and from the Sixth Framework Programme of the European Union via 3DRepertoire (to H.S.). Work in the laboratory of J.M.P. is supported by Boehringer Ingelheim and by Spots of Excellence of the city of Vienna. N.F. is supported by a Boehringer-Ingelheim fellowship. E.W. is supported by a {\textquoteleft}Studienstiftung des deutschen Volkes{\textquoteright} fellowship.",

year = "2008",

month = jan,

doi = "10.1038/nmeth1139",

language = "English",

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pages = "53--55",

journal = "Nature Methods",

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T1 - GraFix

T2 - Sample preparation for single-particle electron cryomicroscopy

AU - Kastner, Berthold

AU - Fischer, Niels

AU - Golas, Monika Mariola

AU - Sander, Bjoern

AU - Dube, Prakash

AU - Boehringer, Daniel

AU - Hartmuth, Klaus

AU - Deckert, Jochen

AU - Hauer, Florian

AU - Wolf, Elmar

AU - Uchtenhagen, Hannes

AU - Urlaub, Henning

AU - Herzog, Franz

AU - Peters, Jan Michael

AU - Poerschke, Dietmar

AU - Lührmann, Reinhard

AU - Stark, Holger

N1 - Funding Information: We thank H. Kohansal, T. Conrad and W. Jahn for expert technical assistance, U. Göringer and M. Brecht for preparing the RNA editing complex, M. Rodnina and K. Gromadski for preparing the 70S ribosome complex, and C. Will for preparing the 17S U2 snRNP. The work was supported by grants from the Federal Ministry of Education and Research, Germany and from the Sixth Framework Programme of the European Union via 3DRepertoire (to H.S.). Work in the laboratory of J.M.P. is supported by Boehringer Ingelheim and by Spots of Excellence of the city of Vienna. N.F. is supported by a Boehringer-Ingelheim fellowship. E.W. is supported by a ‘Studienstiftung des deutschen Volkes’ fellowship.

PY - 2008/1

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GraFix: Sample preparation for single-particle electron cryomicroscopy

Abstract

Forschungsfelder

IMC Forschungsschwerpunkte

ÖFOS 2012 - Österreichischen Systematik der Wissenschaftszweige

UN SDGs

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