MAGMA: Automated assignment of NMR spectra for proteins and complexes
Whilst “traditional” Nuclear Magnetic Resonance (NMR) techniques are able to provide a wealth of information about the structure of small proteins (up to 40 kDa) and their behaviour in solution, more advanced techniques are required for the analysis of larger macromolecules. As 70-80% of eukaryotic proteins assemble into larger structures, the ability to analyse very large proteins is of critical importance in drug discovery. The widespread use of modern techniques, such as methyl TROSY, is inhibited by current data analysis methods that are often time-consuming, error prone and expensive.
Oxford researchers have developed MAGMA, a method for the robust assignment of methyl TROSY spectra. This innovative technology will allow a powerful but under-utilised technique to find a mainstream use for structural biology, drug discovery and agrichemicals.
Probing the structure and dynamic behaviour of proteins and complexes
NMR spectroscopy has become an essential tool for solution-state studies of proteins and other macromolecules. NMR spectroscopy simultaneously provides information about the structure and dynamics of biomolecules at an atomic resolution, thereby enabling analyses of protein folding, denaturation, folding intermediates and transition states, conformational and dynamic behaviour of a biomolecule, ligand binding and mapping of binding sites.
Pushing the limits
Traditional NMR techniques have provided a wealth of information about small proteins, up to approximately 30 kDa in weight. Translating NMR to the study of larger macromolecules has proved challenging. Methyl TROSY (Transverse Relaxation Optimised Spectroscopy) has recently been developed to provide information on the structure and dynamic behaviour of protein complexes up to 1 MDa in weight: capturing 99% of the human proteome. Despite the ability to push beyond the traditional weight limit, the assignment of methyl TROSY spectra is presently time-consuming and cost-prohibitive.
An automated assignment
Currently, assignment of methyl TROSY spectra is typically performed using (multiple rounds of) single point mutagenesis. This requires multiple expensive samples and can provide high cost but potentially unreliable data. Oxford researchers have developed a method that automatically assigns the residues whilst eliminating the need for or enabling targeted (informative) in situ mutagenesis.
Advantages of the Oxford method:
- The graphical user interface supports processing raw data and picking spectral resonances
- Compatible with ‘NMR pipe’ (from NIH) and ‘topspin’ (from Bruker)
- time and cost-effective (105 faster than MAGMA v1)
- measurements taken on a wild-type sample
- provides all feasible solutions, with a confidence fit to direct downstream analyses
- robust data analysis in the presence of artefacts, incomplete detection of resonances, presence of impurities and other sources of experimental error
- 100% accurate
Application of this highly disruptive method removes a key barrier to the widespread adoption of methyl TROSY NMR. Thereby enabling usage of this and related NMR techniques for drug, agrichemical and nutraceutical discovery programmes. By using solution NMR, the Oxford researchers were able to experimentally measure inter-methyl distances from the NOESY data. The method has been extensively tested, and the technique is the subject of a patent application. Oxford University Innovation is seeking commercial partners with an interest in incorporating this technique into their discovery programmes.
NMR Assignment is part of the Oxford Protein Analysis Software Suite (OxPASS), which provides the means to collate data from Mass Spectrometry, NMR and X-Ray crystallography experiments with unprecedented speed and accuracy. Click here for more information.
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