Please use this identifier to cite or link to this item:
http://hdl.handle.net/1942/40281
Title: | Reliability and accuracy of single-molecule FRET studies for characterization of structural dynamics and distances in proteins | Authors: | Agam, Ganesh Gebhardt, Christian Popara, Milana Maechtel, 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 Huebner, 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 Munoz, Gabriel G. Moya Quast, Robert B. Robb, Nicole C. Sattler, Michael Schlierf, Michael Schneider, Jonathan Schroeder, 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 |
Issue Date: | 2023 | Publisher: | NATURE PORTFOLIO | Source: | NATURE METHODS, 20 (4) , p. 523 -535 | Abstract: | 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. | Notes: | Lamb, DC (corresponding author), Ludwig Maximilians Univ Munchen, Dept Chem, Munich, Germany.; Cordes, T (corresponding author), Ludwig Maximilians Univ Munchen, Fac Biol, Phys & Synthet Biol, Planegg Martinsried, Germany.; Barth, A; Seidel, CAM (corresponding author), Heinrich Heine Univ Dusseldorf, Mol Phys Chem, Dusseldorf, Germany.; Barth, A (corresponding author), Delft Univ Technol, Kavli Inst Nanosci, Dept Bionanosci, Delft, Netherlands. a.barth@tudelft.nl; cseidel@hhu.de; d.lamb@lmu.de; cordes@bio.lmu.de |
Document URI: | http://hdl.handle.net/1942/40281 | ISSN: | 1548-7091 | e-ISSN: | 1548-7105 | DOI: | 10.1038/s41592-023-01807-0 | ISI #: | 000961406200002 | Category: | A1 | Type: | Journal Contribution |
Appears in Collections: | Research publications |
Files in This Item:
File | Description | Size | Format | |
---|---|---|---|---|
Reliability and accuracy of single-molecule FRET studies for characterization of structural dynamics and distances in proteins.pdf | Published version | 4.36 MB | Adobe PDF | View/Open |
Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.