The MS/NMR Assay is a protocol for rapidly screening small organic molecules for their ability to bind a target protein while obtaining structure related information as part of a structure based drug discovery and design program. The methodology takes advantage of and combines the inherent strengths of size-exclusion gel chromatography, mass spectrometry and NMR to identify bound complexes in a relatively universal high-throughput screening approach. Size-exclusion gel chromatography in the spin column format provides the high-speed separation of a protein-ligand complex from free ligands. The spin column eluent is then analyzed under denaturing conditions by electrospray ionization (ESI) mass spectrometry (MS) for the presence of small molecular-weight compounds formerly bound to the protein. Hits identified by MS are then individually assayed by chemical shift perturbations in a 2D 1H-15N-HSQC NMR spectrum to verify specific interactions of the compound with the protein and identification of the binding site on the protein.
- R. Powers and M. M. Siegel (2006) Applications of Nuclear Magnetic Resonance and Mass Spectrometry to Anticancer Drug Discovery, Novel Anticancer Agents: Strategies for Discovery and Clinical Testing, A. A. Adjei and J. K. Buolamwini, eds., Elsevier Science, New York, NY, ISBN: 0-12-088561-1.
- Powers, R.; Moy, F. J.; Siegel, M. M.; Mobilio, D. (2001) "Methods of structure-based drug design using MS/NMR", American Home Products Corporation, USA: PCT Int. Appl. US 2001/0046684 A1.
- From: Analytical Chemistry (2001)
- MS vs. NMR for Screening
- Process flow chart of MS/NMR assay and sample preperation robot
- Examples of Gel-filtration-MS Data
- Gel-filtration-MS Titration Results
- Gel-filtration-MS Mixture Data
- Examples of HSQC perturbation data, chemical shift mapping on protein structure and complex structure
- Use MS-NMR without NMR assignments (hypothetical illustration)
- Summary of RGS4 pilot screen
- From: Strategies for Discovery and Clinical Testing of Novel Anticancer Agents. ISBN: 0-12-088561-1.