ArticleProceedings of the National Academy of Sciences of the United States of America2002
Early kinetic intermediate in the folding of acyl-CoA binding protein detected by fluorescence labeling and ultrarapid mixing.
Article in Proceedings of the National Academy of Sciences of the United States of America, 2002. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 31 papers.
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Who cites it
31 citing papers in PubMed.
- Global analysis of thermal and chemical denaturation using CheMelt: Thermodynamic dissection of highly thermostable de novo designed proteins.Protein science : a publication of the Protein Society · 2026Article
- Selective control of ligand binding through a distal mutation that alters the protein native ensemble.Proceedings of the National Academy of Sciences of the United States of America · 2026Article
- Protein structure and folding pathway prediction based on remote homologs recognition using PAthreader.Communications biology · 2023Article
- Advances in Mixer Design and Detection Methods for Kinetics Studies of Macromolecular Folding and Binding on the Microsecond Time Scale.Molecules (Basel, Switzerland) · 2022Article
- Complex Folding Landscape of Apomyoglobin at Acidic pH Revealed by Ultrafast Kinetic Analysis of Core Mutants.The journal of physical chemistry. B · 2018Article
- Unveiling the folding mechanism of the Bromodomains.Biochemistry and biophysics reports · 2017Article
- A J-modulated protonless NMR experiment characterizes the conformational ensemble of the intrinsically disordered protein WIP.Journal of biomolecular NMR · 2016Article
- Protein Ensembles: How Does Nature Harness Thermodynamic Fluctuations for Life? The Diverse Functional Roles of Conformational Ensembles in the Cell.Chemical reviews · 2016Review
- Sandwich-format 3D printed microfluidic mixers: a flexible platform for multi-probe analysis.Journal of micromechanics and microengineering : structures, devices, and systems · 2015Article
- Kinetics of fast changing intramolecular distance distributions obtained by combined analysis of FRET efficiency kinetics and time-resolved FRET equilibrium measurements.Biophysical journal · 2014Article
- Nonuniform chain collapse during early stages of staphylococcal nuclease folding detected by fluorescence resonance energy transfer and ultrarapid mixing methods.Protein science : a publication of the Protein Society · 2013Article
- Slow unfolded-state structuring in Acyl-CoA binding protein folding revealed by simulation and experiment.Journal of the American Chemical Society · 2012Article
- Probing the folding intermediate of Bacillus subtilis RNase P protein by nuclear magnetic resonance.Biochemistry · 2010Article
- Cooperative formation of native-like tertiary contacts in the ensemble of unfolded states of a four-helix protein.Proceedings of the National Academy of Sciences of the United States of America · 2010Article
- Ruggedness in the folding landscape of protein L.HFSP journal · 2008Article
- Barrierless evolution of structure during the submillisecond refolding reaction of a small protein.Proceedings of the National Academy of Sciences of the United States of America · 2008Article
- Folding versus aggregation: polypeptide conformations on competing pathways.Archives of biochemistry and biophysics · 2008Review
- Protein hydrophobic collapse and early folding steps observed in a microfluidic mixer.Biophysical journal · 2007Article
- Folding kinetics of staphylococcal nuclease studied by tryptophan engineering and rapid mixing methods.Journal of molecular biology · 2007Article
- NMR analysis of the conformational properties of the trapped on-pathway folding intermediate of the bacterial immunity protein Im7.Journal of molecular biology · 2007Article
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5 authors.
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Abstract
Early conformational events during folding of acyl-CoA binding protein (ACBP), an 86-residue alpha-helical protein, were explored by using a continuous-flow mixing apparatus with a dead time of 70 micros to measure changes in intrinsic tryptophan fluorescence and tryptophan-dansyl fluorescence energy transfer. Although the folding of ACBP was initially described as a concerted two-state process, the tryptophan fluorescence measurements revealed a previously unresolved phase with a time constant tau = 80 micros, indicating formation of an intermediate with only slightly enhanced fluorescence of Trp-55 and Trp-58 relative to the unfolded state. To amplify this phase, a dansyl fluorophore was introduced at the C terminus by labeling an I86C mutant of ACBP with 5-IAEDANS [5-((((2-iodoacetyl)amino)ethyl)amino)naphthalene-1-sulfonic acid]. Continuous-flow refolding of guanidine HCl-denatured ACBP showed a major increase in tryptophan-dansyl fluorescence energy transfer, indicating formation of a partially collapsed ensemble of states on the 100-micros time scale. A subsequent decrease in dansyl fluorescence is attributed to intramolecular quenching of donor fluorescence on formation of the native state. The kinetic data are fully accounted for by three-state mechanisms with either on- or off-pathway intermediates. The intermediate accumulates to a maximum population of 40%, and its stability depends only weakly on denaturant concentration, which is consistent with a marginally stable ensemble of partially collapsed states with approximately 1/3 of the solvent-accessible surface buried. The findings indicate that ultrafast mixing methods combined with sensitive conformational probes can reveal transient accumulation of intermediate states in proteins with apparent two-state folding mechanisms.
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