| Autor: |
Monaco S; School of Pharmacy, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, U.K., Angulo J; Instituto de Investigaciones Químicas (IIQ), Consejo Superior de Investigaciones Científicas and Universidad de Sevilla, Avenida Américo Vespucio, 49, Sevilla 41092, Spain., Wallace M; School of Pharmacy, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, U.K. |
| Jazyk: |
angličtina |
| Zdroj: |
Journal of the American Chemical Society [J Am Chem Soc] 2023 Aug 02; Vol. 145 (30), pp. 16391-16397. Date of Electronic Publication: 2023 Jul 24. |
| DOI: |
10.1021/jacs.3c02218 |
| Abstrakt: |
We have combined saturation transfer difference NMR (STD NMR) with chemical shift imaging (CSI) and controlled concentration gradients of small molecule ligands to develop imaging STD NMR, a new tool for the assessment of protein-ligand interactions. Our methodology allows the determination of protein-ligand dissociation constants ( K D ) and assessment of the binding specificity in a single NMR tube, avoiding time-consuming titrations. We demonstrate the formation of suitable and reproducible concentration gradients of ligand along the vertical axis of the tube, against homogeneous protein concentration, and present a CSI pulse sequence for the acquisition of STD NMR experiments at different positions along the sample tube. Compared to the conventional methodology in which the [ligand]/[protein] ratio is increased manually, we can perform STD NMR experiments at a greater number of ratios and construct binding epitopes in a fraction (∼20%) of the experimental time. Second, imaging STD NMR also allows us to screen for non-specific binders, by monitoring any variation of the binding epitope map at increasing [ligand]/[protein] ratios. Hence, the proposed method does carry the potential to speed up and smooth out the drug discovery process. |
| Databáze: |
MEDLINE |
| Externí odkaz: |
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