; Gebhardt, Christian ; Popara, Milana
; Mächtel, Rebecca ; Folz, Julian ; Ambrose, Benjamin ; Chamachi, Neharika
; Chung, Sang Yoon
; Craggs, Timothy D.
; de Boer, Marijn ; Grohmann, Dina ; Ha, Taekjip ; Hartmann, Andreas
; Hendrix, Jelle ; Hirschfeld, Verena ; Hübner, Christian G. ; Hugel, Thorsten ; Kammerer, Dominik ; Kang, Hyun-Seo ; Kapanidis, Achillefs N. ; Krainer, Georg ; Kramm, Kevin ; Lemke, Edward A. ; Lerner, Eitan
; Margeat, Emmanuel
; Martens, Kirsten ; Michaelis, Jens
; Mitra, Jaba ; Moya Muñoz, Gabriel G. ; Quast, Robert B.
; Robb, Nicole C. ; Sattler, Michael
; Schlierf, Michael
; Schneider, Jonathan ; Schröder, Tim ; Sefer, Anna ; Tan, Piau Siong ; Thurn, Johann ; Tinnefeld, Philip ; van Noort, John ; Weiss, Shimon ; Wendler, Nicolas ; Zijlstra, Niels ; Barth, Anders
; Seidel, Claus A. M. ; Lamb, Don C. ; Cordes, Thorben 
| Dokumentenart: | Artikel | ||||
|---|---|---|---|---|---|
| Titel eines Journals oder einer Zeitschrift: | Nature Methods | ||||
| Verlag: | NATURE PORTFOLIO | ||||
| Ort der Veröffentlichung: | BERLIN | ||||
| Band: | 20 | ||||
| Nummer des Zeitschriftenheftes oder des Kapitels: | 4 | ||||
| Seitenbereich: | S. 523-535 | ||||
| Datum: | 2023 | ||||
| Institutionen: | Biologie und Vorklinische Medizin > Institut für Biochemie, Genetik und Mikrobiologie > Lehrstuhl für Mikrobiologie (Archaeenzentrum) Biologie und Vorklinische Medizin > Institut für Biochemie, Genetik und Mikrobiologie > Lehrstuhl für Mikrobiologie (Archaeenzentrum) > Prof. Dr. Dina Grohmann | ||||
| Identifikationsnummer: |
| ||||
| Stichwörter / Keywords: | RESONANCE ENERGY-TRANSFER; ALTERNATING-LASER EXCITATION; NANO-POSITIONING SYSTEM; CONFORMATIONAL DYNAMICS; ORIENTATIONAL FREEDOM; LIFETIME DISTRIBUTION; PHOTON DISTRIBUTION; LIGAND-BINDING; RNA; SPECTROSCOPY; | ||||
| Dewey-Dezimal-Klassifikation: | 500 Naturwissenschaften und Mathematik > 550 Geowissenschaften | ||||
| Status: | Veröffentlicht | ||||
| Begutachtet: | Ja, diese Version wurde begutachtet | ||||
| An der Universität Regensburg entstanden: | Ja | ||||
| Dokumenten-ID: | 76294 |
Zusammenfassung
Single-molecule Forster-resonance energy transfer (smFRET) experiments allow the study of biomolecular structure and dynamics in vitro and in vivo. We performed an international blind study involving 19 laboratories to assess the uncertainty of FRET experiments for proteins with respect to the measured FRET efficiency histograms, determination of distances, and the detection and quantification of ...

Zusammenfassung
Single-molecule Forster-resonance energy transfer (smFRET) experiments allow the study of biomolecular structure and dynamics in vitro and in vivo. We performed an international blind study involving 19 laboratories to assess the uncertainty of FRET experiments for proteins with respect to the measured FRET efficiency histograms, determination of distances, and the detection and quantification of structural dynamics. Using two protein systems with distinct conformational changes and dynamics, we obtained an uncertainty of the FRET efficiency <= 0.06, corresponding to an interdye distance precision of <= 2 angstrom and accuracy of <= 5 angstrom. We further discuss the limits for detecting fluctuations in this distance range and how to identify dye perturbations. Our work demonstrates the ability of smFRET experiments to simultaneously measure distances and avoid the averaging of conformational dynamics for realistic protein systems, highlighting its importance in the expanding toolbox of integrative structural biology. An international blind study confirms that smFRET measurements on dynamic proteins are highly reproducible across instruments, analysis procedures and timescales, further highlighting the promise of smFRET for dynamic structural biology.
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