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Measuring translational diffusion of 15N-enriched biomolecules in complex solutions with a simplified 1H-15N HMQC-filtered BEST sequence

  • Shenggen Yao
  • Thomas G. Meikle
  • Ashish Sethi
  • Frances Separovic
  • Jeffrey J. Babon
  • David W. Keizer
Original Article
  • 50 Downloads

Abstract

Pulsed-field gradient nuclear magnetic resonance has seen an increase in applications spanning a broad range of disciplines where molecular translational diffusion properties are of interest. The current study introduces and experimentally evaluates the measurement of translational diffusion coefficients of 15N-enriched biomolecules using a 1H-15N HMQC-filtered band-selective excitation short transient (BEST) sequence as an alternative to the previously described SOFAST-XSTE sequence. The results demonstrate that accurate translational diffusion coefficients of 15N-labelled peptides and proteins can be obtained using this alternative 1H-15N HMQC-filtered BEST sequence which is implementable on NMR spectrometers equipped with probes fitted with a single-axis field gradient, including most cryoprobes dedicated to bio-NMR. The sequence is of potential use for direct quantification of protein or peptide translational diffusion within complex systems, such as in mixtures of macromolecules, crowded solutions, membrane-mimicking media and in bicontinuous cubic phases, where conventional sequences may not be readily applicable due to the presence of intense signals arising from sources other than the protein or peptide under investigation.

Keywords

1H-15N HMQC BEST Complex solutions Crowded solutions Detergent micelles Isotope-filtered PFG-NMR Translational diffusion 

Notes

Acknowledgements

We thank Dr Aitor Moreno of Bruker for the source code of the 1H-15N HSQC-edited PFG-NMR sequence (Fig. S3, Supplemental Materials).

Supplementary material

249_2018_1311_MOESM1_ESM.docx (587 kb)
Supplementary material 1 (DOCX 587 kb)
249_2018_1311_MOESM2_ESM.docx (587 kb)
Supplementary material 2 (DOCX 587 kb)

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Copyright information

© European Biophysical Societies' Association 2018

Authors and Affiliations

  1. 1.Bio21 Molecular Science and Biotechnology InstituteThe University of MelbourneMelbourneAustralia
  2. 2.School of Science, College of Science, Engineering and HealthRMIT UniversityMelbourneAustralia
  3. 3.Department of Biochemistry and Molecular BiologyThe University of MelbourneMelbourneAustralia
  4. 4.School of ChemistryThe University of MelbourneMelbourneAustralia
  5. 5.The Walter and Eliza Hall Institute of Medical ResearchMelbourneAustralia
  6. 6.Department of Medical BiologyThe University of MelbourneMelbourneAustralia

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