ArticleProceedings of the National Academy of Sciences of the United States of America2003
Probing protein oligomerization in living cells with fluorescence fluctuation spectroscopy.
Article in Proceedings of the National Academy of Sciences of the United States of America, 2003. The graph could read no effect estimate from its abstract, so it casts no vote on the map. An erratum has been issued. Cited by 125 papers.
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Who cites it
125 citing papers in PubMed, 212 citations in OpenAlex.
- Protocol to determine oligomerization states of proteins in living cells using photon counting histogram analysis.STAR protocols · 2026Article
- Filament formation and NAD processing by noncanonical human FAM118 sirtuins.Nature structural & molecular biology · 2025Article
- Resolving oligomeric states of photoactivatable proteins in living cells via photon counting histogram analysis.iScience · 2025Article
- The two coiled-coil domains of the fission yeast kinesin-6 Klp9 are required for motor tetramerization and spindle elongation.microPublication biology · 2025Article
- Analysis of Arc/Arg3.1 Oligomerization In Vitro and in Living Cells.International journal of molecular sciences · 2024Article
- Statistical analysis of the autocorrelation function in fluorescence correlation spectroscopy.Biophysical journal · 2024Article
- Determining the Functional Oligomeric State of Membrane-Associated Protein Oligomers Forming Membrane Pores on Giant Lipid Vesicles.Analytical chemistry · 2023Article
- Autocorrelation function of finite-length data in fluorescence correlation spectroscopy.Biophysical journal · 2023Article
- Benchmarking of novel green fluorescent proteins for the quantification of protein oligomerization in living cells.PloS one · 2023Article
- The dependence of EGFR oligomerization on environment and structure: A camera-based N&B study.Biophysical journal · 2022Article
- Article
- Radial pair correlation of molecular brightness fluctuations maps protein diffusion as a function of oligomeric state within live-cell nuclear architecture.Biophysical journal · 2022Article
- Combined FCS and PCH Analysis to Quantify Protein Dimerization in Living Cells.International journal of molecular sciences · 2021Article
- Single-molecule microscopy for in-cell quantification of protein oligomeric stoichiometry.Current opinion in structural biology · 2021Review
- Gain-of-Function Properties of a Dynamin 2 Mutant Implicated in Charcot-Marie-Tooth Disease.Frontiers in cellular neuroscience · 2021Article
- Number and Brightness Analysis: Visualization of Protein Oligomeric State in Living Cells.Advances in experimental medicine and biology · 2021Article
- Agonist binding directs dynamic competition among nuclear receptors for heterodimerization with retinoid X receptor.The Journal of biological chemistry · 2020Article
- Differentiating Luminal and Membrane-Associated Nuclear Envelope Proteins.Biophysical journal · 2020Article
- Monomeric cohesin state revealed by live-cell single-molecule spectroscopy.EMBO reports · 2020Article
- Identifying Heteroprotein Complexes in the Nuclear Envelope.Biophysical journal · 2020Article
65 more citing papers are in PubMed but not listed here.
Corrections and comments
- Erratum issued
Authors and funding
3 authors at 1 institution in 1 country.
Funding
Abstract
Fluorescence fluctuation spectroscopy provides information about protein interactions in the intercellular environment from naturally occurring equilibrium fluctuations. We determine the molecular brightness of fluorescent proteins from the fluctuations by analyzing the photon counting histogram (PCH) or its moments and demonstrate the use of molecular brightness in probing the oligomerization state of proteins. We report fluorescence fluctuation measurements of enhanced GFP (EGFP) in cells up to concentrations of 10 microM by using an improved PCH theory. The molecular brightness of EGFP is constant in the concentration range studied. The brightness of a tandem EGFP construct, which carries two fluorophores, increases by a factor of two compared with EGFP alone, demonstrating the sensitivity of molecular brightness as a probe for protein complex formation. Oligomerization of nuclear receptors plays a crucial role in the regulation of gene expression. We probe the oligomerization state of the testicular receptor 4 and the ligand-binding domains of retinoid X receptor and retinoic acid receptor by observing molecular brightness changes as a function of protein concentration. The large concentration range accessible by experiment allows us to perform titration experiments on EGFP fusion proteins. An increase in the molecular brightness with protein concentration indicates the formation of homocomplexes. We observe the formation of homodimers of retinoid X receptor ligand binding domain upon addition of ligand. Resolving protein interactions in a cell is an important step in understanding cellular function on a molecular level. Brightness analysis promises to develop into an important tool for determining protein complex formation in cells.
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