Characterisation of Photoaffinity‐Based Chemical Probes by Fluorescence Imaging and Native‐State Mass Spectrometry

Teruya, Kanae; Rankin, Gregory M.; Chrysanthopoulos, Panagiotis K.; Tonissen, Kathryn Fay; Poulsen, Sally‐Ann

doi:10.1002/cbic.201600598

Cited by 6 publications

(8 citation statements)

References 24 publications

(20 reference statements)

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“…The interactions between proteins and ligands are crucial to proper cellular function. , The structures, functions, and interactions of protein–ligand complexes can be significantly affected by salts. − Specific metal ion cofactors can regulate the bioactivity of proteins . In native mass spectrometry (MS), ligand–protein interactions are normally stabilized using volatile salts at high ionic strengths to rapidly and directly measure the mass, binding stoichiometry, and relative ligand–protein binding affinities with high sensitivity. − However, most biochemical approaches to probe protein–ligand interactions, including nuclear magnetic resonance spectroscopy, circular dichroism spectroscopy, isothermal titration calorimetry, and optical spectroscopy, use nonvolatile salts that can more accurately reflect the in vivo environment of the protein–ligand complex. However, nonvolatile salts and common biological buffers readily adduct to proteins ions to result in broad spectral peaks that have deleterious effects on mass spectra by lowering the sensitivity and signal-to-noise ratios and increasing background chemical noise .…”

Section: Introductionmentioning

confidence: 99%

Nanoscale Ion Emitters in Native Mass Spectrometry for Measuring Ligand–Protein Binding Affinities

et al. 2019

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Electrospray ionization (ESI) mass spectrometry (MS) is a crucial method for rapidly determining the interactions between small molecules and proteins with ultrahigh sensitivity. However, nonvolatile molecules and salts that are often necessary to stabilize the native structures of protein–ligand complexes can readily adduct to protein ions, broaden spectral peaks, and lower signal-to-noise ratios in native MS. ESI emitters with narrow tip diameters (∼250 nm) were used to significantly reduce the extent of adduction of salt and nonvolatile molecules to protein complexes to more accurately measure ligand–protein binding constants than by use of conventional larger-bore emitters under these conditions. As a result of decreased salt adduction, peaks corresponding to protein–ligand complexes that differ in relative molecular weight by as low as 0.06% can be readily resolved. For low-molecular-weight anion ligands formed from sodium salts, anion-bound and unbound protein ions that differ in relative mass by 0.2% were completely baseline resolved using nanoscale emitters, which was not possible under these conditions using conventional emitters. Owing to the improved spectral resolution obtained using narrow-bore emitters and an analytically derived equation, K d values were simultaneously obtained for at least six ligands to a single druggable protein target from one spectrum for the first time. This research suggests that ligand–protein binding constants can be directly and accurately measured from solutions with high concentrations of nonvolatile buffers and salts by native MS.

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Section: Introductionmentioning

confidence: 99%

Nanoscale Ion Emitters in Native Mass Spectrometry for Measuring Ligand–Protein Binding Affinities

et al. 2019

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“…Poulsen started by describing the complementarity of MS alongside other more commonly used methods to detect fragment binding . She then explained her progress using MS to identify a new zinc binding chemotype and strategy to use MS for SAR . MS emerges as an exciting alternative for FBDD.…”

Section: Day 2: the Sweet Spot In Chemical Biologymentioning

confidence: 99%

“…33 She then explained her progress using MS to identify a new zinc binding chemotype 34 and strategy to use MS for SAR. 35 MS emerges as an exciting alternative for FBDD.…”

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confidence: 99%

Meeting Proceedings from ICBS 2018—Toward Translational Impact

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“…Carbonic anhydrase (CA, E.C.4.2.1.1) isoforms are zinc metalloenzymes found in a miscellany of organisms and mainly associated with catalyzing the foundational reaction, CO 2 molecule hydration into bicarbonate and protons . The inhibitors of carbonic anhydrase isoenzymes (CAIs) are used in therapeutic applications in many diseases .…”

Section: Introductionmentioning

confidence: 99%

New Isoindole‐1,3‐dione Substituted Sulfonamides as Potent Inhibitors of Carbonic Anhydrase and Acetylcholinesterase: Design, Synthesis, and Biological Evaluation

et al. 2019

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Herein, a series of isoindole‐1,3‐dione substituted sulfonamide derivatives (3, 4 a–k) were designed, synthesized, and biologically evaluated, as inhibitors of carbonic anhydrase (CA) and acetylcholinesterase (AChE). CA and AChE inhibitory activities of newly synthesized isoindole‐1,3‐dione substituted sulfonamides compounds (3, 4 a–k) towards the hCA I, II, and AChE were evaluated utilizing the Verpoorte's and Ellman's assays and checked against that of standard inhibitors, acetazolamide (AAZ) and tacrine (TAC). The developed compounds (3, 4 a–k) showed the potent hCA isoenzyme inhibitory effect with Ki constants ranging from 7.96 to 48.34 nM, compared to AAZ (Kis; 436.20 nM for hCA I and 93.53 nM for hCA II). Among these derivatives; 1,3‐dioxo‐1,3‐dihydroisobenzofuran‐5‐carbocyclic acid (3) and benzyl‐1,3‐dioxo‐2‐(4‐sulfomophenyl)isoindoline‐5‐carboxylate (4 i) determined to be effective AChE inhibitors (Kis, 103.51 and 108.92 nM, respectively); these compounds were almost as potent to TAC (Ki, 109.75 nM). Furthermore, molecular docking studies of derivatives 3 and 4 i were carried out utilizing the crystal structures of hCA I (PDB Code: 4WR7), II (PDB Code: 4HT0) isozymes and AChE (PDB Code: 4EY7) receptors to study their binding interactions.

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Characterisation of Photoaffinity‐Based Chemical Probes by Fluorescence Imaging and Native‐State Mass Spectrometry

Cited by 6 publications

References 24 publications

Nanoscale Ion Emitters in Native Mass Spectrometry for Measuring Ligand–Protein Binding Affinities

Nanoscale Ion Emitters in Native Mass Spectrometry for Measuring Ligand–Protein Binding Affinities

Meeting Proceedings from ICBS 2018—Toward Translational Impact

New Isoindole‐1,3‐dione Substituted Sulfonamides as Potent Inhibitors of Carbonic Anhydrase and Acetylcholinesterase: Design, Synthesis, and Biological Evaluation

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