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15N CPMG Relaxation Dispersion for the Investigation of Protein Conformational Dynamics on the µs-ms Timescale
Published on: April 19, 2021
Multiplet-filtered and gradient-selected zero-quantum TROSY experiments for 13C1H3 methyl groups in proteins
Michelle L Gill1, Arthur G Palmer
1Department of Biochemistry and Molecular Biophysics, Columbia University, 630 West 168th Street, New York, NY 10032, USA.
Journal of Biomolecular NMR
|September 16, 2011
Summary
New NMR experiments improve the measurement of protein structures by better suppressing unwanted signals. These gradient-selected heteronuclear zero-quantum coherence (gsHZQC) TROSY methods enhance accuracy for methyl groups in proteins.
Area of Science:
- Biophysics
- Structural Biology
- Nuclear Magnetic Resonance (NMR) Spectroscopy
Background:
- Heteronuclear zero-quantum coherence (HZQC) experiments are vital for protein structure determination.
- Existing HZQC sequences struggle with suppressing outer multiplet components in medium-sized or flexible proteins.
- Improved signal suppression is crucial for accurate analysis of protein dynamics and structure.
Purpose of the Study:
- To develop and describe novel multiplet-filtered and gradient-selected heteronuclear zero-quantum coherence (gsHZQC) TROSY experiments.
- To enhance the measurement of (1)H-(13)C correlations specifically for (13)CH(3) methyl groups in proteins.
- To provide a method with improved suppression of undesirable spectral components compared to existing HZQC techniques.
Main Methods:
- Implementation of multiplet-filtered and gradient-selected HZQC TROSY pulse sequences.
- Utilized Hahn-echo versions for measuring zero- and double-quantum transverse relaxation rates.
- Applied the developed sequences to Escherichia coli ribonuclease HI (18 kD) for validation.
Main Results:
- The gsHZQC experiments demonstrated improved suppression of the heteronuclear zero-quantum multiplet's outer components.
- Effective application was shown in medium-sized proteins and flexible regions of larger proteins.
- The Hahn-echo versions enabled the identification of chemical exchange broadening via relaxation rate measurements.
- Validation on E. coli ribonuclease HI confirmed enhanced multiplet suppression without significant sensitivity loss.
Conclusions:
- The developed gsHZQC TROSY experiments offer superior performance for (1)H-(13)C correlation measurements in methyl groups.
- These methods provide enhanced spectral clarity, aiding in the structural and dynamic analysis of proteins.
- The technique is valuable for studying proteins where conventional HZQC methods face limitations, such as medium-sized or flexible systems.
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