A5017487591- Advanced Fluorescence Microscopy Techniques
- Protein Structure and Dynamics
- Advanced Biosensing Techniques and Applications
- Biotin and Related Studies
- Alzheimer's disease research and treatments
- Advanced Electron Microscopy Techniques and Applications
- Enzyme Structure and Function
- Monoclonal and Polyclonal Antibodies Research
- Spectroscopy and Quantum Chemical Studies
- Photosynthetic Processes and Mechanisms
- RNA and protein synthesis mechanisms
- DNA and Nucleic Acid Chemistry
- Supramolecular Self-Assembly in Materials
- Metabolomics and Mass Spectrometry Studies
- Plant nutrient uptake and metabolism
- Lanthanide and Transition Metal Complexes
- Plant Molecular Biology Research
- Lipid Membrane Structure and Behavior
- Advanced NMR Techniques and Applications
- Cell Adhesion Molecules Research
- Bioinformatics and Genomic Networks
- HIV Research and Treatment
- HER2/EGFR in Cancer Research
- Computational Drug Discovery Methods
- Prion Diseases and Protein Misfolding
National Institutes of Health
2018-2024
National Institute of Diabetes and Digestive and Kidney Diseases
2018-2024
Korea Advanced Institute of Science and Technology
2011-2016
Yonsei University
2016
Institute for Basic Science
2016
Co-immunoprecipitation (co-IP) has become a standard technique, but its protein-band output provides only static, qualitative information about protein-protein interactions. Here we demonstrate real-time single-molecule co-IP technique that generates videos of individual interactions as they occur in unpurified cell extracts. By analysing single Ras-Raf with 50-ms time resolution, have observed transient intermediates the interaction and determined all essential kinetic rates. Using this...
The bHLH transcription factor PHYTOCHROME INTERACTING FACTOR1 (PIF1) binds G-box elements in vitro and inhibits light-dependent germination Arabidopsis thaliana. A previous genome-wide analysis of PIF1 targeting indicated that 748 sites imbibed seeds, only 59% which possess elements. This suggests the is not sole determinant targeting. to specific could be stabilized by PIF1-interacting factors (PTFs) bind other nearby sequence Here, we report are enriched with G-boxes but also hexameric...
Abstract Intrinsically disordered proteins (IDPs) usually fold during binding to target proteins. In contrast interactions between folded proteins, this additional folding step makes the process more complex. Understanding mechanism of coupled and IDPs requires analysis pathways that involve formation transient complex (TC). However, experimental characterization TC is challenging because it only appears for a very brief period binding. Here, we use single-molecule fluorescence spectroscopy...
Significance There are various diseases caused by protein aggregation such as Alzheimer’s, Parkinson’s, and Huntington’s diseases. From the diversity in fibril structure, is expected to occur via heterogeneous pathways. However, characterization of this heterogeneity extremely difficult because it requires following individual formation a mixture from early oligomerization stages. In work, we investigated 42-residue isoform amyloid β (Aβ42) using single-molecule fluorescence imaging deep...
Abstract We describe theory, experiments, and analyses of three-color Förster resonance energy transfer (FRET) spectroscopy for probing sub-millisecond conformational dynamics protein folding binding disordered proteins. devise a scheme that uses single continuous-wave laser excitation the donor instead alternating one acceptors. This alleviates photophysical problems acceptors such as rapid photobleaching, which is crucial high time resolution experiments with elevated illumination...
We describe the theory, experiment, and analysis of three-color Förster resonance energy transfer (FRET) spectroscopy for probing conformational dynamics a fast-folding protein, α3D. In FRET, site-specific labeling fluorophores is required to avoid ambiguity resulting from various species with different combinations positions. To this end, we first attached two dyes cysteine residue an unnatural amino acid then appended C-terminus protein by sortase-mediated ligation attaching third dye....
Abstract T-box riboswitches are multi-domain noncoding RNAs that surveil individual amino acid availabilities in most Gram-positive bacteria. T-boxes directly bind specific tRNAs, query their aminoacylation status to detect starvation, and feedback control the transcription or translation of downstream amino-acid metabolic genes. Most rapidly recruit cognate tRNA ligands through an intricate three-way stem I-stem II-tRNA interaction, whose establishment is not understood. Using...
Extremely weak protein–protein interactions (PPIs), signified by micromolar or even millimolar dissociation constants, are one of the keys to understanding rapid responses cellular systems. Although single-molecule methods particularly useful in determining kinetics biological processes, their application is largely limited rather strong because diffraction-limited observation volume. In this study, we report a method that allows characterization PPIs using prey concentration 4 orders...
Significance The cosolvent effect alters the equilibrium between native and unfolded states of a protein, with denaturant cosolutes shifting toward latter osmolyte stabilizing former. Quantitative characterization strength cosolute–protein interactions at atomic resolution is experimentally challenging as these are highly transient occur low occupancy. Here, we make use paramagnetic NMR solvent relaxation enhancement measurements to quantify interaction drkN SH3, metastable protein domain...
Abstract Protein aggregation is implicated as the cause of pathology in various diseases such Alzheimer’s and Parkinson’s disease. Polymorphism structure fibrils formed by suggests existence many different assembly pathways therefore a heterogeneous ensemble soluble oligomers. Characterization this heterogeneity key to understanding mechanism toxicity specific oligomers, but practice it extremely difficult because oligomers cannot be readily separated. Here, we investigate highly...
The conventional co-immunoprecipitation provides static and qualitative information about protein-protein interactions. Lee et al. report real-time imaging of process with single-molecule resolution, allowing for characterization the native Ras proteins derived from individual cancers.