Abstract
We describe a (19)F NMR method for detecting bromodomain-ligand interactions using fluorine-labeled aromatic amino acids due to the conservation of aromatic residues in the bromodomain binding site. We test the sensitivity, accuracy, and speed of this method with small molecule ligands (+)-JQ1, BI2536, Dinaciclib, TG101348, and acetaminophen using three bromodomains Brd4, BrdT, and BPTF. Simplified (19)F NMR spectra allowed for simultaneous testing of multiple bromodomains to assess selectivity and identification of a new BPTF ligand. Fluorine labeling only modestly affected the Brd4 structure and function assessed by isothermal titration calorimetry, circular dichroism, and X-ray crystallography. The speed, ease of interpretation, and low concentration of protein needed for binding experiments affords a new method to discover and characterize both native and new ligands.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, Non-P.H.S.
MeSH terms
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Acetaminophen / chemistry
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Amino Acids, Aromatic / chemistry*
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Antigens, Nuclear / chemistry
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Antigens, Nuclear / genetics
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Azepines / chemistry
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Binding Sites
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Bridged Bicyclo Compounds, Heterocyclic / chemistry
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Cell Cycle Proteins
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Crystallography, X-Ray
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Cyclic N-Oxides
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Escherichia coli / genetics
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Escherichia coli / metabolism
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Gene Expression
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Halogenation
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Humans
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Indolizines
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Ligands
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Magnetic Resonance Spectroscopy / methods
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Molecular Docking Simulation
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Molecular Probes / chemistry*
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Nerve Tissue Proteins / chemistry
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Nerve Tissue Proteins / genetics
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Nuclear Proteins / chemistry
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Nuclear Proteins / genetics
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Oncogene Proteins, Fusion / chemistry*
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Oncogene Proteins, Fusion / genetics
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Protein Binding
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Pteridines / chemistry
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Pyridinium Compounds / chemistry
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Pyrrolidines / chemistry
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Recombinant Proteins / chemistry
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Recombinant Proteins / genetics
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Staining and Labeling / methods
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Sulfonamides / chemistry
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Transcription Factors / chemistry
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Transcription Factors / genetics
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Triazoles / chemistry
Substances
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(+)-JQ1 compound
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Amino Acids, Aromatic
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Antigens, Nuclear
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Azepines
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BI 2536
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BRD4 protein, human
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BRDT protein, human
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Bridged Bicyclo Compounds, Heterocyclic
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Cell Cycle Proteins
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Cyclic N-Oxides
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Indolizines
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Ligands
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Molecular Probes
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Nerve Tissue Proteins
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Nuclear Proteins
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Oncogene Proteins, Fusion
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Pteridines
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Pyridinium Compounds
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Pyrrolidines
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Recombinant Proteins
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Sulfonamides
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Transcription Factors
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Triazoles
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fetal Alzheimer antigen
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Acetaminophen
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dinaciclib
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fedratinib