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Related Concept Videos

¹H NMR: Interpreting Distorted and Overlapping Signals01:02

¹H NMR: Interpreting Distorted and Overlapping Signals

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Spin systems where the difference in chemical shifts of the coupled nuclei is greater than ten times J are called first-order spin systems. These nuclei are weakly coupled, and their chemical shifts and coupling constant can generally be estimated from the well-separated signals in the spectrum.
As Δν decreases and the signals move closer, the doublets appear increasingly distorted. The intensities of the inner lines increase at the cost of those of the outer lines as the signals are...
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¹H NMR: Complex Splitting01:13

¹H NMR: Complex Splitting

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A proton M that is coupled to a proton X results in doublet signals for M. However, NMR-active nuclei can be simultaneously coupled to more than one nonequivalent nucleus. When M is coupled to a second proton A, such as in styrene oxide, each peak in the doublet is split into another doublet.
Splitting diagrams or splitting tree diagrams are routinely used to depict such complex couplings. While drawing splitting diagrams, the splitting with the larger coupling constant is usually applied...
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¹³C NMR: ¹H–¹³C Decoupling01:04

¹³C NMR: ¹H–¹³C Decoupling

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The probability of having two carbon-13 atoms next to each other is negligible because of the low natural abundance of carbon-13. Consequently, peak splitting due to carbon-carbon spin-spin coupling is not observed in spectra. However, protons up to three sigma bonds away split the carbon signal according to the n+1 rule, resulting in complicated spectra.
A broadband decoupling technique is used to simplify these complex, sometimes overlapping, signals. Broadband decoupling relies on a...
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¹H NMR of Labile Protons: Deuterium (²H) Substitution00:48

¹H NMR of Labile Protons: Deuterium (²H) Substitution

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This lesson illustrates the role of deuterium substitution in simplifying the NMR spectrum of compounds comprising labile protons. One method employed is the use of deuterium. Amongst the three isotopes of hydrogen, deuterium (2H) has a nucleus composed of one proton and one neutron. When the D2O solvent is added to a pure dry ethanol solution, its labile proton is substituted with deuterium.
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¹H NMR of Labile Protons: Temporal Resolution01:10

¹H NMR of Labile Protons: Temporal Resolution

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Protons bonded to heteroatoms such as nitrogen and oxygen exhibit a range of chemical shift values. This is due to the varying degree of hydrogen bonding between the proton and the heteroatom in other molecules. The extent of hydrogen bonding affects the electron density around the proton, thereby giving different chemical shift values for the protons in the proton NMR spectrum.
The –OH proton in alcohols typically appears in the range of δ 2 to 5 ppm but can vary depending on the specific...
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Mass Spectrometry: Amine Fragmentation00:55

Mass Spectrometry: Amine Fragmentation

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Amines can be identified using mass spectroscopy based on their characteristic fragmentation patterns. The molecular ions of amines undergo fragmentation via ⍺-cleavage. The ⍺-cleavage of the carbon-carbon bonds in amines generates an alkyl radical and resonance-stabilized nitrogen-containing cation.
In amines, the number of nitrogen atoms affects the mass of the molecular ion, which is described by the nitrogen rule of mass spectrometry. This rule states that a compound containing a single...
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Dissolution Dynamic Nuclear Polarization Instrumentation for Real-time Enzymatic Reaction Rate Measurements by NMR
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Label-free NMR-based dissociation kinetics determination.

Pablo Trigo-Mouriño1, Christian Griesinger1, Donghan Lee2

  • 1Department of NMR-Based Structural Biology, Max-Planck Institute for Biophysical Chemistry, Göttingen, Germany.

Journal of Biomolecular NMR
|November 17, 2017
PubMed
Summary

This study introduces a novel high-power relaxation dispersion (RD) method to measure weak binding ligand dissociation rates from receptors without isotopic labeling. This technique offers a direct and efficient way to determine off-rates for drug development.

Keywords:
Ligand bindingNMRRelaxation dispersion

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Area of Science:

  • Biochemistry
  • Chemical Biology
  • Pharmacology

Background:

  • Understanding molecular dissociation is crucial for modulating biomedical interactions.
  • Optimizing drug dissociation rates impacts efficacy, selectivity, and safety.
  • Accurate determination of ligand-receptor off-rates is essential for drug design.

Purpose of the Study:

  • To present a new application of high-power relaxation dispersion (RD) for determining dissociation rates of weak binding ligands.
  • To demonstrate a method that avoids isotopic labeling and simplifies detection.
  • To enable direct access to binding off-rates without serial sample analysis.

Main Methods:

  • Utilized high-power relaxation dispersion (RD) spectroscopy.
  • Probed proton relaxation dispersion on the ligand, eliminating the need for isotopic labeling.
  • Employed large ligand excess and large spin-lock fields for enhanced detection sensitivity and broader rate access.

Main Results:

  • Successfully determined dissociation rates for weak binding ligands.
  • The method proved effective for small molecule interactions with macromolecules like bovine serum albumin and tubulin heterodimers.
  • Achieved detection of faster dissociation rates compared to other relaxation approaches.

Conclusions:

  • The high-power RD method provides a direct, efficient, and label-free approach to measure ligand-receptor dissociation rates.
  • This technique simplifies the experimental process and expands the range of measurable off-rates.
  • The findings have significant implications for optimizing drug candidates by precisely characterizing their binding kinetics.