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Precision and accuracy of single-molecule FRET measurements—a multi-laboratory benchmark study
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(2018) Precision and accuracy of single-molecule FRET measurements—a multi-laboratory benchmark study. Nature Methods, 15 (9). pp. 669-676. doi:10.1038/s41592-018-0085-0 ISSN 1548-7091.
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Official URL: http://dx.doi.org/10.1038/s41592-018-0085-0
Abstract
Single-molecule Förster resonance energy transfer (smFRET) is increasingly being used to determine distances, structures, and dynamics of biomolecules in vitro and in vivo. However, generalized protocols and FRET standards to ensure the reproducibility and accuracy of measurements of FRET efficiencies are currently lacking. Here we report the results of a comparative blind study in which 20 labs determined the FRET efficiencies (E) of several dye-labeled DNA duplexes. Using a unified, straightforward method, we obtained FRET efficiencies with s.d. between ±0.02 and ±0.05. We suggest experimental and computational procedures for converting FRET efficiencies into accurate distances, and discuss potential uncertainties in the experiment and the modeling. Our quantitative assessment of the reproducibility of intensity-based smFRET measurements and a unified correction procedure represents an important step toward the validation of distance networks, with the ultimate aim of achieving reliable structural models of biomolecular systems by smFRET-based hybrid methods.
| Item Type: | Journal Article | ||||||||||||||||||||||||||||||||||||
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| Subjects: | Q Science > QC Physics Q Science > QP Physiology |
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| Divisions: | Faculty of Science, Engineering and Medicine > Medicine > Warwick Medical School > Biomedical Sciences > Cell & Developmental Biology Faculty of Science, Engineering and Medicine > Medicine > Warwick Medical School > Biomedical Sciences Faculty of Science, Engineering and Medicine > Medicine > Warwick Medical School |
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| Library of Congress Subject Headings (LCSH): | Energy transfer, Energy transfer -- Measurement, Molecular dynamics | ||||||||||||||||||||||||||||||||||||
| Journal or Publication Title: | Nature Methods | ||||||||||||||||||||||||||||||||||||
| Publisher: | Nature Publishing Group | ||||||||||||||||||||||||||||||||||||
| ISSN: | 1548-7091 | ||||||||||||||||||||||||||||||||||||
| Official Date: | September 2018 | ||||||||||||||||||||||||||||||||||||
| Dates: |
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| Volume: | 15 | ||||||||||||||||||||||||||||||||||||
| Number: | 9 | ||||||||||||||||||||||||||||||||||||
| Page Range: | pp. 669-676 | ||||||||||||||||||||||||||||||||||||
| DOI: | 10.1038/s41592-018-0085-0 | ||||||||||||||||||||||||||||||||||||
| Status: | Peer Reviewed | ||||||||||||||||||||||||||||||||||||
| Publication Status: | Published | ||||||||||||||||||||||||||||||||||||
| Access rights to Published version: | Open Access (Creative Commons) | ||||||||||||||||||||||||||||||||||||
| Date of first compliant deposit: | 28 July 2020 | ||||||||||||||||||||||||||||||||||||
| Date of first compliant Open Access: | 29 July 2020 | ||||||||||||||||||||||||||||||||||||
| RIOXX Funder/Project Grant: |
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