The first automated 470.59 MHz19F NMR-controlled titration: dissociation constants and ion-specific chemical shifts of 2-amino-4-fluoro 2-methylpent-4-enoic acid

2002 ◽  
Vol 40 (9) ◽  
pp. 573-580 ◽  
Author(s):  
C. E. Uhlemann ◽  
C. G. Pfaff ◽  
G. Hägele ◽  
K. W. Laue ◽  
M. Lübke ◽  
...  
Keyword(s):  
Dissociation Constants ◽  
Chemical Shifts ◽  
Enoic Acid ◽  
Specific Chemical

scholarly journals Protolysis and Complex Formation of Organophosphorus Compounds—Characterization by NMR-Controlled Titrations

Molecules ◽  
2019 ◽  
Vol 24 (18) ◽  
pp. 3238 ◽  
Author(s):  
Hägele
Keyword(s):  
Dissociation Constants ◽  
Organophosphorus Compounds ◽  
Chemical Shifts ◽  
2D Nmr ◽  
Coupling Constants ◽  
Phosphonic Acids ◽  
Specific Chemical ◽  
Hyphenated Technique ◽  
Aminophosphonic Acids ◽  
2D Nmr Spectra

Phosphonic acids, aminophosphonic acids, and phosphonocarboxylic acids are characterized by an advanced hyphenated technique, combining potentiometric titration with NMR spectroscopy. Automated measurements involving 13C, 19F and 31P nuclei lead to “pseudo 2D NMR” spectra, where chemical shifts or coupling constants are correlated with analytical parameters. Dissociation constants, stability constants, dynamic and specific chemical shifts are determined. Macroscopic and microscopic dissociation equilibria are discussed.

scholarly journals Direct assessment of substrate binding to the Neurotransmitter:Sodium Symporter LeuT by solid state NMR

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
Simon Erlendsson ◽  
Kamil Gotfryd ◽  
Flemming Hofmann Larsen ◽  
Jonas Sigurd Mortensen ◽  
Michel-Andreas Geiger ◽  
...  
Keyword(s):  
Solid State ◽  
Binding Site ◽  
Solid State Nmr ◽  
Chemical Shifts ◽  
Substrate Binding ◽  
Experimental Conditions ◽  
Specific Chemical ◽  
Sodium Dependent ◽  
Number Of Binding Sites ◽  
Neurotransmitter:Sodium Symporter

The Neurotransmitter:Sodium Symporters (NSSs) represent an important class of proteins mediating sodium-dependent uptake of neurotransmitters from the extracellular space. The substrate binding stoichiometry of the bacterial NSS protein, LeuT, and thus the principal transport mechanism, has been heavily debated. Here we used solid state NMR to specifically characterize the bound leucine ligand and probe the number of binding sites in LeuT. We were able to produce high-quality NMR spectra of substrate bound to microcrystalline LeuT samples and identify one set of sodium-dependent substrate-specific chemical shifts. Furthermore, our data show that the binding site mutants F253A and L400S, which probe the major S1 binding site and the proposed S2 binding site, respectively, retain sodium-dependent substrate binding in the S1 site similar to the wild-type protein. We conclude that under our experimental conditions there is only one detectable leucine molecule bound to LeuT.

scholarly journals Formation of highly quaternized N,N,N-trimethylchitosan: a chemoselective methodology in aqueous media

2019 ◽  
Vol 91 (3) ◽  
pp. 489-496 ◽  
Author(s):  
Youssouf D. Soubaneh ◽  
Steeven Ouellet ◽  
Caroline Dion ◽  
Jonathan Gagnon
Keyword(s):  
Biomedical Applications ◽  
Chemical Shifts ◽  
Aqueous Media ◽  
Acid Base ◽  
Specific Chemical ◽  
H Nmr ◽  
Low Degree ◽  
Simple Step ◽  
Carbonate Salt

Abstract N,N,N-Trimethylchitosan (TMC) represents a rare example of cationic polysaccharides and numerous studies have shown its potential in biological and biomedical applications. TMC with high degrees of quaternization (DQ) were synthesized from N-methylation of N,N-dimethylchitosan (DMC), which was obtained by reductive alkylation of high molecular weight chitosan in a simple step process and in good yields. The effects of base and solvents were evaluated on the quaternization reaction. The N-methylation of DMC was performed selectively by CH3I and carbonate in water where quaternization was achieved quantitatively with a low degree of O-methylation (17 %). Moreover, the greener procedure allows easy recovery and purification by conventional filtration as a carbonate salt, in which the anion can be exchanged by an acid-base reaction. Quantification of DQ involving 1H NMR integration of methyl peaks must be performed on protonated TMC. High field NMR spectra of TMC showed two specific chemical shifts for anomeric peaks (5.0 and 5.4 ppm) that can also be used for the determination of DQ. This latter method avoids the superimposition problems with other pyranosyl peaks.

Asymmetric protonation of glutamate residues drives a preferred transport pathway in EmrE

2021 ◽  
Vol 118 (41) ◽  
pp. e2110790118
Author(s):  
Jianping Li ◽  
Ampon Sae Her ◽  
Nathaniel J. Traaseth
Keyword(s):  
Lipid Bilayers ◽  
Efflux Pump ◽  
Dissociation Constants ◽  
Chemical Shifts ◽  
Acid Dissociation ◽  
Coupled Transport ◽  
Wild Type ◽  
Multidrug Efflux Pump ◽  
Transport Pathway ◽  
Small Multidrug Resistance

EmrE is an Escherichia coli multidrug efflux pump and member of the small multidrug resistance (SMR) family that transports drugs as a homodimer by harnessing energy from the proton motive force. SMR family transporters contain a conserved glutamate residue in transmembrane 1 (Glu14 in EmrE) that is required for binding protons and drugs. Yet the mechanism underlying proton-coupled transport by the two glutamate residues in the dimer remains unresolved. Here, we used NMR spectroscopy to determine acid dissociation constants (pKa) for wild-type EmrE and heterodimers containing one or two Glu14 residues in the dimer. For wild-type EmrE, we measured chemical shifts of the carboxyl side chain of Glu14 using solid-state NMR in lipid bilayers and obtained unambiguous evidence on the existence of asymmetric protonation states. Subsequent measurements of pKa values for heterodimers with a single Glu14 residue showed no significant differences from heterodimers with two Glu14 residues, supporting a model where the two Glu14 residues have independent pKa values and are not electrostatically coupled. These insights support a transport pathway with well-defined protonation states in each monomer of the dimer, including a preferred cytoplasmic-facing state where Glu14 is deprotonated in monomer A and protonated in monomer B under pH conditions in the cytoplasm of E. coli. Our findings also lead to a model, hop-free exchange, which proposes how exchangers with conformation-dependent pKa values reduce proton leakage. This model is relevant to the SMR family and transporters comprised of inverted repeat domains.

scholarly journals Interact: Automated analysis of protein-ligand interactions by 1D and 2D NMR

2018 ◽  
Author(s):  
Pierre Millard ◽  
Guy Lippens
Keyword(s):  
Dissociation Constants ◽  
Chemical Shifts ◽  
Automated Analysis ◽  
2D Nmr ◽  
Nmr Titration ◽  
Protein Ligand Interactions ◽  
Fitting Procedures ◽  
Ligand Interactions ◽  
Titration Data ◽  
Automated Tools

AbstractNMR titration experiments contain rich information on the thermodynamic, kinetic and structural aspects of protein-ligand interactions. Automated tools are required to process the large number of signals typically acquired in these experiments and facilitate quantitative interpretations. We present Interact, a Python script accessible within the Bruker BioSpin TopSpin™ software, which allows automated analysis of both 1D and 2D NMR titration experiments. Interact performs peak picking and annotation of the successive spectra and supports quantitative interpretation of changes in chemical shifts and linewidths induced by the ligand (e.g. to estimate dissociation constants) through different fitting procedures. Interact can be applied to all types of 1D and 2D NMR experiments and all nuclei, hence facilitating routine analysis of existing and forthcoming NMR titration data. Interact was implemented in Python and can be used on Windows, Unix and MacOS platforms. The source code is distributed under OpenSource license at http://github.com/MetaSys-LISBP/Interact.

Preparation and nuclear magnetic resonance studies of diaryloxytriphenylphosphoranes. Correlation of substituent effects with 31P N.M.R. chemical shifts

1984 ◽  
Vol 37 (12) ◽  
pp. 2447 ◽  
Author(s):  
Mv Itzstein ◽  
ID Jenkins
Keyword(s):  
Nuclear Magnetic Resonance ◽  
Dissociation Constants ◽  
Chemical Shifts ◽  
Substituent Effects ◽  
Acid Dissociation ◽  
Linear Free Energy Relationship ◽  
Acid Dissociation Constants ◽  
Magnetic Resonance Studies ◽  
Linear Free Energy ◽  
Electron Donating Groups

Triphenylphosphine and diisopropyl azodicarboxylate react with phenols in tetrahydrofuran or chloroform at 0° to give diaryloxytriphenylphosphoranes. A linear free-energy relationship has been found between the 31P n.m.r. chemical shifts of many of these phosphoranes and the acid dissociation constants of the corresponding phenols. In general, electron-withdrawing groups on the phenol result in downfield 31P n.m.r. chemical shifts, while electron donating groups result in up-field shifts.

Interaction of troponin I and troponin C: 19F NMR studies of the binding of the inhibitory troponin I peptide to turkey skeletal troponin C

1991 ◽  
Vol 69 (9) ◽  
pp. 674-681 ◽  
Author(s):  
A. Patricia Campbell ◽  
Paul J. Cachia ◽  
Brian D. Sykes
Keyword(s):  
Calcium Binding ◽  
Troponin I ◽  
Dissociation Constants ◽  
Chemical Shifts ◽  
Troponin C ◽  
Resonance Spectroscopy ◽  
Nmr Studies ◽  
Binding Model ◽  
Peptide Binding Site ◽  
Titration Data

We have used 19F nuclear magnetic resonance spectroscopy to study the interaction of the inhibitory region of troponin (TnI) with apo- and calcium(II)-saturated turkey skeletal troponin C (TnC), using the synthetic TnI analogue Nα-acetyl[19FPhe106]TnI(104–115)amide. Dissociation constants of Kd = (3.7 ± 3.1) × 10−5 M for the apo interaction and Kd = (4.8 ± 1.8) × 10−5 M for the calcium(II)-saturated interaction were obtained using a 1:1 binding model of peptide to protein. The 19F NMR chemical shifts for the F-phenylalanine of the bound peptide are different from the apo- and calcium-saturated protein, indicating a different environment for the bound peptide. The possibility of 2:1 binding of the peptide to Ca(II)-saturated TnC was tested by calculating the fit of the experimental titration data to a series of theoretical binding curves in which the dissociation constants for the two hypothetical binding sites were varied. We obtained the best fit for 0.056 mM ≤ Kd1 ≤ 0.071 mM and 0.5 mM ≤ Kd2 ≤ 2.0 mM. These results allow the possibility of a second peptide binding site on calcium(II)-saturated TnC with an affinity 10- to 20-fold weaker than that of the first site.Key words: tropinin C, tropinin I, calcium binding, NMR studies, muscle proteins.

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