Assignment of 19F Resonances in Protein Solution State NMR Studies

Kitevski-LeBlanc, Julianne L.; Prosser, R. Scott

doi:10.1007/978-3-642-16712-6_314

Cited by 15 publications

(21 citation statements)

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“…19 F NMR spectroscopy has a long history of application for biomolecular characterization, with three key advantages: (i) high sensitivity and wide chemical shift range; (ii) an almost complete absence in biological systems, leading to a lack of background signals; and, (iii) the potential to employ 1D experiments, allowing for decreased experiment times relative to triple‐resonance experiments . In triple‐resonance studies, chemical shift overlap in large proteins may prove intractable, even with sparse labeling .…”

Section: Figurementioning

confidence: 99%

“…The lack of 19 F chemical shift perturbation implies that neither ligand changes the local electronic environment significantly. This is not trivially interpretable since, at present, the relative contributions from the various effects underlying 19 F chemical shifts remain controversial . Given favorable SDS micelle binding by both apela‐32 and apelin‐36, one potential reason for HSQC spectral perturbation would be nonspecific binding of a given ligand to the micelle, indirectly modulating AR structure and/or dynamics.…”

Section: Figurementioning

confidence: 99%

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Mixed Fluorotryptophan Substitutions at the Same Residue Expand the Versatility of ¹⁹F Protein NMR Spectroscopy

Kenward

Shin

Rainey

2018

Chemistry A European J

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The strategy of applying fluorine NMR to characterize ligand binding to a membrane protein prepared with mixtures of tryptophans substituted with F at different positions on the indole ring was tested. The F NMR behavior of 4-, 5-, 6-, and 7-fluorotryptophan were directly compared as a function of both micellar environment and fragment size for two overlapping apelin receptor (AR/APJ) segments; one with a single transmembrane (TM) helix and two tryptophan residues, the other with three TM helices and two additional tryptophan residues. Chemical shifts, peak patterns, and nuclear spin relaxation rates were observed to vary as a function of micellar conditions and F substitution position in the indole ring, with the exposure of a given residue to micelle or solvent being the primary differentiating factor. Titration of the 3-TM AR segment biosynthetically prepared as a mixture of 5- and 7-fluorotryptophan-containing isoforms by two distinct peptide ligands (apelin-36 and apela-32) demonstrated site-specific F peak intensity changes for one ligand but not the other. In contrast, both ligands perturbed H- N HSQC peak patterns to a similar degree. Characterization of multiple fluorotryptophan types for a given set of tryptophan residues, thus, significantly augments the potential to apply F NMR to track otherwise obscure modulation of protein conformation and dynamics without an explicit requirement for mutagenesis or chemical modification.

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Section: Figurementioning

confidence: 99%

Section: Figurementioning

confidence: 99%

Mixed Fluorotryptophan Substitutions at the Same Residue Expand the Versatility of ¹⁹F Protein NMR Spectroscopy

Kenward

Shin

Rainey

2018

Chemistry A European J

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“…Naturally occurring proteins do not contain any fluorine nuclei. However, synthetically fluorinated amino acid analogues can be incorporated into proteins . Incorporation of fluorinated amino acid residue requires a bacterial strain for recombinant protein expression, which is auxotrophic for a given amino acid.…”

Section: Protein‐based Methodsmentioning

confidence: 99%

“…Cysteines are favorable residues due to their ability to form covalent bonds through their side chain sulfhydryl group. However the side chain NH 2 of lysine or hydroxyl group of serine/threonine can also be used as a possible site of reaction . In order to investigate protein–ligand interactions by 19 F NMR a ligand titration using a series of simple 1D fluorine spectra has to be recorded.…”

Section: Protein‐based Methodsmentioning

confidence: 99%

Investigating Protein–Ligand Interactions by Solution Nuclear Magnetic Resonance Spectroscopy

et al. 2018

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Protein–ligand interactions are of fundamental importance in almost all processes in living organisms. The ligands comprise small molecules, drugs or biological macromolecules and their interaction strength varies over several orders of magnitude. Solution NMR spectroscopy offers a large repertoire of techniques to study such complexes. Here, we give an overview of the different NMR approaches available. The information they provide ranges from the simple information about the presence of binding or epitope mapping to the complete 3 D structure of the complex. NMR spectroscopy is particularly useful for the study of weak interactions and for the screening of binding ligands with atomic resolution.

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“…In particular, 19 F diffusion‐ordered spectroscopy ( 19 F DOSY) is a rapidly developing approach that can provide valuable information on the molecular sizes in small molecules and polymers . Typically, fluorinated moieties are introduced into proteins by expression of a protein with a non‐canonical fluorine‐containing amino acid, or by a cysteine functionalization . Incorporation of fluorine labels into peptides can be made by applying fluorinated amino acid building blocks during solid‐phase peptide synthesis or, alternatively, by residue modification of the peptide on‐resin .…”

Section: Methodsmentioning

confidence: 99%

Selective ¹⁹F‐Labeling of Functionalized Carboxylic Acids with Difluoromethyl Diazomethane (CF₂HCHN₂)

Mykhailiuk

Kishko

Kubyshkin

et al. 2017

Chemistry A European J

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The incorporation of fluorinated moieties into organic structures is a particularly powerful approach for generating compounds with useful biological applications in industry, pharmacology, and agriculture. Here, the recently developed reagent-CF HCHN -selectively esterifies carboxyl groups on organic molecules in the presence of other polar functional groups. The reaction is remarkably simple, proceeding at room temperature in chloroform without addition of a catalyst. The procedure allows "last-step" incorporation of a fluorinated ester moiety into organic structures, including bioorganic molecules such as peptides and proteins. The fluorinated ester can subsequently function as a F-probe in biomolecules, thereby enabling molecular size determination by diffusion F NMR experiments.

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Assignment of 19F Resonances in Protein Solution State NMR Studies

Cited by 15 publications

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Mixed Fluorotryptophan Substitutions at the Same Residue Expand the Versatility of ¹⁹F Protein NMR Spectroscopy

Mixed Fluorotryptophan Substitutions at the Same Residue Expand the Versatility of ¹⁹F Protein NMR Spectroscopy

Investigating Protein–Ligand Interactions by Solution Nuclear Magnetic Resonance Spectroscopy

Selective ¹⁹F‐Labeling of Functionalized Carboxylic Acids with Difluoromethyl Diazomethane (CF₂HCHN₂)

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Assignment of 19F Resonances in Protein Solution State NMR Studies

Cited by 15 publications

References 0 publications

Mixed Fluorotryptophan Substitutions at the Same Residue Expand the Versatility of 19F Protein NMR Spectroscopy

Mixed Fluorotryptophan Substitutions at the Same Residue Expand the Versatility of 19F Protein NMR Spectroscopy

Investigating Protein–Ligand Interactions by Solution Nuclear Magnetic Resonance Spectroscopy

Selective 19F‐Labeling of Functionalized Carboxylic Acids with Difluoromethyl Diazomethane (CF2HCHN2)

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Mixed Fluorotryptophan Substitutions at the Same Residue Expand the Versatility of ¹⁹F Protein NMR Spectroscopy

Mixed Fluorotryptophan Substitutions at the Same Residue Expand the Versatility of ¹⁹F Protein NMR Spectroscopy

Selective ¹⁹F‐Labeling of Functionalized Carboxylic Acids with Difluoromethyl Diazomethane (CF₂HCHN₂)