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Related Experiment Video

Updated: Jul 26, 2026

Frequency Mixing Magnetic Detection Scanner for Imaging Magnetic Particles in Planar Samples

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Published on: June 9, 2016

How to detect internal motion by homonuclear NMR.

Jürgen Schleucher¹, Sybren S Wijmenga

¹Department of Medical Biochemistry and Biophysics, and Department of Organic Chemistry, Umeå University, S-90187 Umeå, Sweden. jurgen@rabbit.chem.umu.se

Journal of the American Chemical Society

|May 16, 2002

Summary

This study introduces an off-resonance ROESY method to measure molecular internal motions by analyzing cross-peak intensities. The technique reveals detailed insights into protein dynamics, applicable to molecules up to 15 kDa without isotope labeling.

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Area of Science:

Structural Biology
Biophysical Chemistry
Nuclear Magnetic Resonance Spectroscopy

Background:

Nuclear Overhauser Effect Spectroscopy (NOESY) and Rotating-frame Overhauser Effect Spectroscopy (ROESY) cross-peak intensities are influenced by internuclear distances and molecular motion.
Internal molecular motion is often disregarded, with NOESY data solely interpreted for internuclear distances.

Purpose of the Study:

To develop and validate a method using off-resonance ROESY to quantify molecular internal motions.
To assess the utility of this method for analyzing protein dynamics and its applicability to molecules up to 15 kDa.

Main Methods:

Utilized off-resonance ROESY experiments to measure a weighted average of NOE and ROE, controlled by an angle theta.
Developed a protocol to measure theta;(0) from a series of off-resonance ROESY spectra, analyzing individual cross-peaks to evaluate internal motions of specific H,H vectors.
Applied the method to the protein BPTI, analyzing 75 cross-peaks to determine theta;(0) values and their precision.

Main Results:

The off-resonance ROESY method successfully determined theta;(0) values, reflecting both overall and internal molecular motion.
Internal motions on a 100-300 ps timescale showed the largest reduction in theta;(0).
Analysis of BPTI revealed well-defined theta;(0) for rigid regions and identified dynamics in flexible regions and methyl groups, consistent with known timescales.

Conclusions:

Off-resonance ROESY provides a sensitive method for monitoring internal molecular motions, particularly useful for H,H contacts in structure calculations.
The technique is applicable to molecules up to approximately 15 kDa and does not require isotope labeling, broadening its use to natural products.
The method offers precise measurements of molecular dynamics, complementing existing structural biology techniques.