ArticleProtein science : a publication of the Protein Society2012
Simplified proteomics approach to discover protein-ligand interactions.
Article in Protein science : a publication of the Protein Society, 2012. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 23 papers.
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23 citing papers in PubMed, 56 citations in OpenAlex.
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- Comparative Analysis of Protein Folding Stability-Based Profiling Methods for Characterization of Biological Phenotypes.Journal of the American Society for Mass Spectrometry · 2023Article
- Toward the analysis of functional proteoforms using mass spectrometry-based stability proteomics.Frontiers in analytical science · 2023Article
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- Chemoproteomic-enabled characterization of small GTPase Rab1a as a target of anRSC chemical biology · 2022Article
- Metabolization and sequestration of plant specialized metabolites in insect herbivores: Current and emerging approaches.Frontiers in physiology · 2022Review
- Osteosarcoma cell proliferation suppression via SHP-2-mediated inactivation of the JAK/STAT3 pathway by tubocapsenolide A.Journal of advanced research · 2021Article
- Analysis of Brain Protein Stability Changes in Mouse Models of Normal Aging and α-Synucleinopathy Reveals Age- and Disease-Related Differences.Journal of proteome research · 2021Article
- Protein Folding Stability Changes Across the Proteome Reveal Targets of Cu Toxicity inACS chemical biology · 2021Article
- Chaperone action of a cofactor in protein folding.Protein science : a publication of the Protein Society · 2020Article
- Comparative Analysis of Mass-Spectrometry-Based Proteomic Methods for Protein Target Discovery Using a One-Pot Approach.Journal of the American Society for Mass Spectrometry · 2020Article
- Chemo-Selection Strategy for Limited Proteolysis Experiments on the Proteomic Scale.Analytical chemistry · 2018Article
- Proteome-Wide Structural Biology: An Emerging Field for the Structural Analysis of Proteins on the Proteomic Scale.Journal of proteome research · 2018Review
- Energetic Coupling between Ligand Binding and Dimerization in Escherichia coli Phosphoglycerate Mutase.Biochemistry · 2016Article
- Characterization of the Saccharomyces cerevisiae ATP-Interactome using the iTRAQ-SPROX Technique.Journal of the American Society for Mass Spectrometry · 2016Article
- Global analysis of protein folding thermodynamics for disease state characterization.Journal of proteome research · 2015Article
- SILAC-pulse proteolysis: A mass spectrometry-based method for discovery and cross-validation in proteome-wide studies of ligand binding.Journal of the American Society for Mass Spectrometry · 2014Article
- StableIsotope Labeling with Amino Acids in Cell Culture (SILAC)-based strategy for proteome-wide thermodynamic analysis of protein-ligand binding interactions.Molecular & cellular proteomics : MCP · 2014Article
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4 authors at 1 institution in 1 country.
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Abstract
Identifying targets of biologically active small molecules is an essential but still challenging task in drug research and chemical genetics. Energetics-based target identification is an approach that utilizes the change in the conformational stabilities of proteins upon ligand binding in order to identify target proteins. Different from traditional affinity-based capture approaches, energetics-based methods do not require any labeling or immobilization of the test molecule. Here, we report a surprisingly simple version of energetics-based target identification, which only requires ion exchange chromatography, SDS PAGE, and minimal use of mass spectrometry. The complexity of a proteome is reduced through fractionation by ion exchange chromatography. Urea-induced unfolding of proteins in each fraction is then monitored by the significant increase in proteolytic susceptibility upon unfolding in the presence and the absence of a ligand. Proteins showing a different degree of unfolding with the ligand are identified by SDS PAGE followed by mass spectrometry. Using this approach, we identified ATP-binding proteins in the Escherichia coli proteome. In addition to known ATP-binding proteins, we also identified a number of proteins that were not previously known to interact with ATP. To validate one such finding, we cloned and purified phosphoglyceromutase, which was not previously known to bind ATP, and confirmed that ATP indeed stabilizes this protein. The combination of fractionation and pulse proteolysis offers an opportunity to investigate protein-drug or protein-metabolite interactions on a proteomic scale with minimal instrumentation and without modification of a molecule of interest.
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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.