Identification of Differential Protein Binding Affinities in an Atropisomeric Pharmaceutical Compound by Noncovalent Mass Spectrometry, Equilibrium Dialysis, and Nuclear Magnetic Resonance
| Autor: | Matthew P. Crump, Harry Mackenzie, Christine E. Prosser, Adam Hold, John Crosby, Daniel James Ford, John Robert Porter, Ian Whitcombe, Richard J. K. Taylor, Rachel A. Garlish, Hannah J. Maple |
|---|---|
| Rok vydání: | 2013 |
| Předmět: |
Atropisomer
Magnetic Resonance Spectroscopy Chemistry Drug discovery Stereochemistry bcl-X Protein Plasma protein binding Mass spectrometry Combinatorial chemistry Affinities Mass Spectrometry Analytical Chemistry Pharmaceutical Preparations Proto-Oncogene Proteins c-bcl-2 Antiapoptotic protein Equilibrium dialysis Chirality (chemistry) Protein Binding |
| Zdroj: | Analytical Chemistry. 85:5958-5964 |
| ISSN: | 1520-6882 0003-2700 |
| Popis: | Atropisomerism of pharmaceutical compounds is a challenging area for drug discovery programs (Angew. Chem., Int. Ed. 2009, 48, 6398-6401). Strategies for dealing with these compounds include raising the energy barrier to atropisomerization in order to develop the drug as a single isomer (Tetrahedron 2004, 60, 4337-4347) or reducing the barrier to rotation and developing a mixture of rapidly interconverting isomers (Chirality 1996, 8, 364-371). Commonly, however, the atropisomers will be differentiated in terms of their affinity for a given protein target, and it is therefore important to rapidly identify the most active component prior to further compound development. We present equilibrium dialysis and saturation transfer difference NMR (STD-NMR) as techniques for assessing relative affinities of an atropisomeric mixture against antiapoptotic protein targets Bcl-2 and Bcl-xL. These techniques require no prior separation of the mixture of compounds and are therefore rapid and simple approaches. We also explore the use of noncovalent mass spectrometry for determining KD values of individual atropisomers separated from the equilibrium mixture and compare the results to solution-phase measurements. Results from equilibrium dialysis, STD-NMR, and noncovalent mass spectrometry are all in excellent agreement and provide complementary information on differential binding, amplification of the strongest binders, and KD values. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|