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Related Concept Videos

2D NMR: Heteronuclear Single-Quantum Correlation Spectroscopy (HSQC)01:19

2D NMR: Heteronuclear Single-Quantum Correlation Spectroscopy (HSQC)

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Heteronuclear single-quantum correlation spectroscopy (HSQC) is a 2D NMR technique that reveals one-bond correlations between hydrogen and a heteronucleus. The HSQC experiment is similar to the heteronuclear correlation experiment (HETCOR) but is more sensitive. In the HSQC spectrum, the proton chemical shift is plotted on the horizontal F2 axis, while the 13C chemical shift is plotted on the vertical F1 axis. The corresponding proton and 13C spectra are also shown. The HSQC contour plot does...
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¹H NMR of Conformationally Flexible Molecules: Temporal Resolution00:52

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At room temperature, the chair conformer of cyclohexane undergoes rapid ring flipping between two equivalent chair conformers at a rate of approximately 105 times per second. These two chair conformers are in equilibrium. The rapid ring flipping results in the interconversion of the axial proton to an equatorial proton and an equatorial to the axial proton. Such interconversions are too rapid and cannot be detected on the NMR timescale. Hence, the NMR spectrometer cannot distinguish between the...
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Intrinsically Disordered Proteins02:18

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Intrinsically disordered proteins are a group of proteins that do not fold into specific three-dimensional structures. Their structural flexibility allows them to complement ordered proteins to perform functions that are inaccessible to rigid structures. They are more common in eukaryotes than prokaryotes and may either be exclusively intrinsically disordered or hybrid proteins, consisting of a mix of ordered and disordered regions. The absence of a rigid structure in these proteins can be...
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¹H NMR of Conformationally Flexible Molecules: Variable-Temperature NMR01:15

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The axial and equatorial protons in cyclohexane can be distinguished by performing a variable-temperature NMR experiment. In this process, except for one proton, the remaining eleven protons are replaced by deuterium. The deuterium substitution avoids the possible peak splitting caused by the spin-spin coupling between the adjacent protons. The remaining proton flips between the axial and equatorial positions.
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¹H NMR of Labile Protons: Deuterium (²H) Substitution00:48

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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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Related Experiment Video

Updated: Apr 7, 2026

Analyzing Protein Dynamics Using Hydrogen Exchange Mass Spectrometry
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Transient Helicity in the Intrinsically Disordered Protein ACTR Measured by Hydrogen Exchange.

I Simina Cuciurean1, Christian Buch Parsbæk1, Kasper D Rand2

  • 1Structural Biology and NMR Laboratory, Department of Biology, University of Copenhagen, 2200 Copenhagen, Denmark.

Biochemistry
|April 6, 2026
PubMed
Summary
This summary is machine-generated.

Hydrogen exchange, measured by NMR and MS, effectively detects subtle helical changes in intrinsically disordered proteins (IDPs). This method complements NMR chemical shifts for studying protein dynamics and transient structures.

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A Hydrogen-Deuterium Exchange Mass Spectrometry HDX-MS Platform for Investigating Peptide Biosynthetic Enzymes
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Area of Science:

  • Biochemistry
  • Structural Biology
  • Biophysics

Background:

  • Intrinsically disordered proteins (IDPs) are crucial for cellular processes, often utilizing dynamic structures for function.
  • Nuclear Magnetic Resonance (NMR) chemical shifts are standard for IDP secondary structure analysis.
  • Hydrogen exchange (HX) is a valuable tool for folded proteins, but its application to IDPs requires further validation.

Purpose of the Study:

  • To systematically evaluate hydrogen exchange (HX) methods for detecting transient helicity in intrinsically disordered proteins (IDPs).
  • To compare the sensitivity of HX measurements with NMR chemical shifts in IDP variants.
  • To establish HX as a reliable technique for characterizing dynamic protein structures.

Main Methods:

  • Utilized Hydrogen exchange measured by NMR and MS (HDX-MS) on four variants of the ACTR activation domain with varying helical propensities.
  • Developed pseudo-protection factors derived from NMR-based exchange rates for robust inter-variant comparison.
  • Correlated HX-derived pseudo-protection factors with helicity information from NMR chemical shifts.

Main Results:

  • Hydrogen exchange measurements showed strong correlation with helicity determined by NMR chemical shifts.
  • Pseudo-protection factors effectively resolved subtle differences in transient helicity among IDP variants.
  • Demonstrated the sensitivity and reproducibility of HDX-MS for characterizing dynamic protein structures.

Conclusions:

  • Hydrogen exchange is a sensitive and reproducible method for characterizing transient structures in intrinsically disordered proteins.
  • HDX-MS serves as a valuable complementary technique to NMR chemical shift analysis for IDPs.
  • This study validates HX as a powerful tool for probing local structure and dynamics in IDPs.