Exploring weak ligand-protein interactions by long-lived NMR states: improved contrast in fragment-based drug screening.
Bottom Line: In this work, we describe the use of LLS for competitive binding experiments to measure accurate dissociation constants of fragments that bind weakly to the ATP binding site of the N-terminal ATPase domain of heat shock protein 90 (Hsp90), a therapeutic target for cancer treatment.The LLS approach allows one to characterize ligands with an exceptionally wide range of affinities, since it can be used for ligand concentrations [L] that are several orders of magnitude smaller than the dissociation constants K(D).This property makes the LLS method particularly attractive for the initial steps of fragment-based drug screening, where small molecular fragments that bind weakly to a target protein must be identified, which is a difficult task for many other biophysical methods.
Affiliation: Institut des Sciences et Ingénierie Chimiques, Ecole Polytechnique Fédérale de Lausanne, Batochime (BCH), 1015 Lausanne (Switzerland). email@example.com.Show MeSH
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Mentions: In the absence of competing binders, the interaction between the spy ligand and the protein leads to rapid LLS relaxation and hence to the attenuation of the LLS signal (spectrum 1 in Figure 4); conversely, the presence of a competitor leads to a partial displacement of the spy ligand, hence to slower LLS relaxation and a partial restoration of the LLS signal of the spy (spectrum 2 in Figure 4). This change in LLS signal is due to a mere 13 % change in the amount of bound ligand, which itself is only 0.3 % of the total ligand concentration.
Affiliation: Institut des Sciences et Ingénierie Chimiques, Ecole Polytechnique Fédérale de Lausanne, Batochime (BCH), 1015 Lausanne (Switzerland). firstname.lastname@example.org.