[NMR] Protein structure determination using NMR spectroscopy

[NMR] Protein structure determination using NMR spectroscopy

Protein structure determination using Nuclear Magnetic Resonance (NMR) spectroscopy is one of the three core techniques in Structural Biology. This module will provide an in-depth introduction to the technique. No similar module is offered at this level anywhere in South Africa.


Dr David Pugh (Deputy Programme Co-ordinator, UWC) has a background in Applied Mathematics and Physics, and has experience of determining protein structures using nuclear magnetic resonance (NMR) spectroscopy. Dr Pugh is the leader of the Structural Biology Group at UWC. He will also co-ordinate the NMR component of the Masters Programme in Structural Biology, as well as contribute lectures on mathematical techniques and NMR spectroscopy.

Main Outcomes

The aim of this module is to introduce you to the practice and some of the theory of protein structure determination using Nuclear Magnetic Resonance Spectroscopy. By the end you should be able to:

  • understand the principles of NMR spectroscopy,
  • be able to use appropriate software to process and analyse spectra,
  • be able to analyse one, two and three-dimensional spectra in order to assign chemical shifts and NOE’s,
  • interpret NOE’s as spatial restraints between atoms
  • be able to use molecular dynamics calculations to determine protein structures using NOE-derived distance restraints.
  • use NMR-based methods for analyzing protein-protein interactions and the dynamical behaviour of proteins in solution
  • use NMR to measure substrate binding to proteins

Main Content

  • Principles of biomolecular NMR spectroscopy
  • Spectral processing
  • Multi-dimensional experiments
  • Chemical shift assignment
  • Structure determination
  • NMR-based interaction studies
  • Backbone dynamics

This module descriptor document also can be downloaded as an MS-Word document.

Home Department: Biotechnology, UWC
Module description (Header): Protein NMR spectroscopy
Generic module name: Structural Biology
Alpha-numeric code: STB705
Credit Value: 15 Credits
Duration: 6 Weeks
Module Type: P
Level: 8
Prerequisites: Computational and Mathematical Primer, Principles of Protein Structure,
Structural Bioinformatics and Molecular Modelling
Co-requisites: None
Prohibited combinations: None
Learning time breakdown (hours):
Contact with lecturer/tutor: 60
Assignments & tasks: 40
Tests & examinations: 5
Practicals: 0
Selfstudy: 35
Project: 10
Total Learning Time 150
Methods of Student Assessment: Tutorial sheets will be handed out approximately once a week. Tutorials will count approximately equally towards a tutorial mark, which will make up 40% of the final mark for the module. A 3-hour exam will be scheduled at the end of the course.
40% Tutorials
20% Project
40% Exam


  1. Week 1 Introduction to Biomolecular NMR
  2. Week 2 Spectral processing
    1. Fourier Transformation of a Lorentzian signal
    2. NMRPipe and NMRDraw
    3. Phasing
    4. Window functions
    5. Spectral width; the Nyquist theorem; folding
    6. Referencing
    7. Tutorial: Processing of HSQC spectrum
  3. Week 3 Spectral assignment
    1. NMRView
    2. Working with 3D datasets; strip plots
    3. Tutorial: Working with HNCO, HNCA, HN(CO)CA spectra
    4. Tutorial: Backbone assignment using CBCA(CO)NH
    5. Side-chain assignment using HCCH-TOCSY and HSQC-TOCSY
  4. Week 4 Structure determination
    1. Aria-CNS
    2. Molmol
  5. Week 1-3 1-D NMR
    1. acquire 1-D spectrum of protein
    2. vector model of NMR, coherence, T1 and T2
    3. data processing, window functions, sweep width, resolution
    4. saturation, pulsed field gradients
    5. software tools
    6. data simulations and analysis using MATLAB
  6. Week 4-5 2-D NMR
    1. coherence transfer
    3. assignment of spin systems
  7. Week 6 Heteronuclear experiments
    1. isotope enrichment
    2. 15N-separated NOESY, TOCSY
    3. inter-residue coherence transfer experiments, HNCA, HN(CO)CA
  8. Week 7-8 Structure calculations
    1. chemical shifts and secondary structure
    2. generation of spatial restraints
    3. calculations using CNS, Dyanna

Online Lectures

Introduction to Biomolecular NMR


Claridge, TDW. 2000. High-Resolution NMR Techniques in Organic Chemistry. Amsterdam: Pergamon, Tetrahedron Organic Chemistry, Vol. 19.

Ernst, RR, G Bodenhausen, and A Wokaun. 1990. Principles of Nuclear Magnetic Resonance in One and Two Dimensions. Oxford, UK: Clarendon Press, International Series of Monographs on Chemistry, Vol. 14.

Homans, SW. 1992. A Dictionary of Concepts in NMR. Clarendon Press, Biophysical Techniques Series, Vol. 1, Revised Edition.

James, TL. 1998. "Fundamentals of NMR." In Biophysics Textbook Online,, Bethesda, MD: Biophysical Society, http://www.biophysics.org/btol/.

Leach, AR. 2001. Molecular Modelling: Principles and Applications. Harlow, England: Prentice Hall, 2nd ed.

Lerner, L and DA Horita. 1993. "Teaching high-resolution nuclear magnetic resonance to graduate students in biophysics." Biophysical Journal, 65 (6): 2692-2697.

Levitt, MH. 2008. Spin Dynamics: Basics of Nuclear Magnetic Resonance. Chichester: Wiley, 2nd edition.