ArticleBiophysical journal2010
Distinguishing between protein dynamics and dye photophysics in single-molecule FRET experiments.
Article in Biophysical journal, 2010. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 29 papers.
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Who cites it
29 citing papers in PubMed, 60 citations in OpenAlex.
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- Modification of Cysteine-Substituted Antibodies Using Enzymatic Oxidative Coupling Reactions.Bioconjugate chemistry · 2023Article
- Labeling of Proteins for Single-Molecule Fluorescence Spectroscopy.Methods in molecular biology (Clifton, N.J.) · 2022Article
- Secondary modification of oxidatively-modified proline N-termini for the construction of complex bioconjugates.Organic & biomolecular chemistry · 2020Article
- Diverse Folding Pathways of HIV-1 Protease Monomer on a Rugged Energy Landscape.Biophysical journal · 2019Article
- Ensemble and single-molecule FRET studies of protein synthesis.Methods (San Diego, Calif.) · 2018Article
- Effects of Fluorophore Attachment on Protein Conformation and Dynamics Studied by spFRET and NMR Spectroscopy.Chemistry (Weinheim an der Bergstrasse, Germany) · 2017Article
- Oligomerization of the tetramerization domain of p53 probed by two- and three-color single-molecule FRET.Proceedings of the National Academy of Sciences of the United States of America · 2017Article
- Förster resonance energy transfer: Role of diffusion of fluorophore orientation and separation in observed shifts of FRET efficiency.PloS one · 2017Article
- Analysis of Fluorescence Lifetime and Energy Transfer Efficiency in Single-Molecule Photon Trajectories of Fast-Folding Proteins.The journal of physical chemistry. B · 2016Article
- A Method for Extracting the Free Energy Surface and Conformational Dynamics of Fast-Folding Proteins from Single Molecule Photon Trajectories.The journal of physical chemistry. B · 2015Article
- Movement of elongation factor G between compact and extended conformations.Journal of molecular biology · 2015Article
- A starting point for fluorescence-based single-molecule measurements in biomolecular research.Molecules (Basel, Switzerland) · 2014Review
- Single-Molecule Analysis of CytochromeThe journal of physical chemistry letters · 2013Article
- Article
- Combined versatile high-resolution optical tweezers and single-molecule fluorescence microscopy.The Review of scientific instruments · 2012Article
- Nanochannel-based single molecule recycling.Nano letters · 2012Article
- Observation of protein folding/unfolding dynamics of ubiquitin trapped in agarose gel by single-molecule FRET.European biophysics journal : EBJ · 2012Article
- Exploring one-state downhill protein folding in single molecules.Proceedings of the National Academy of Sciences of the United States of America · 2012Article
- Photophysics of fluorescent probes for single-molecule biophysics and super-resolution imaging.Annual review of physical chemistry · 2012Review
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Authors and funding
3 authors at 1 institution in 1 country.
Funding
Abstract
Förster resonance energy transfer (FRET) efficiency distributions in single-molecule experiments contain both structural and dynamical information. Extraction of this information from these distributions requires a careful analysis of contributions from dye photophysics. To investigate how mechanisms other than FRET affect the distributions obtained by counting donor and acceptor photons, we have measured single-molecule fluorescence trajectories of a small alpha/beta protein, i.e., protein GB1, undergoing two-state, folding/unfolding transitions. Alexa 488 donor and Alexa 594 acceptor dyes were attached to cysteines at positions 10 and 57 to yield two isomers-donor(10)/acceptor(57) and donor(57)/acceptor(10)-which could not be separated in the purification. The protein was immobilized via binding of a histidine tag added to a linker sequence at the N-terminus to cupric ions embedded in a polyethylene-glycol-coated glass surface. The distribution of FRET efficiencies assembled from the trajectories is complex with widths for the individual peaks in large excess of that caused by shot noise. Most of this complexity can be explained by two interfering photophysical effects-a photoinduced red shift of the donor dye and differences in the quantum yield of the acceptor dye for the two isomers resulting from differences in quenching rate by the cupric ion. Measurements of steady-state polarization, calculation of the donor-acceptor cross-correlation function from photon trajectories, and comparison of the single molecule and ensemble kinetics all indicate that conformational distributions and dynamics do not contribute to the complexity.
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