Determination of Ligand-Binding Affinity (Kd) Using Transverse Relaxation Rate (R2) in the Ligand-Observed 1H NMR Experiment and Applications to Fragment-Based Drug Discovery.
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Authors
Liu, M
Mirza, A
McAndrew, PC
Thapaliya, A
Pierrat, OA
Stubbs, M
Hahner, T
Chessum, NEA
Innocenti, P
Caldwell, J
Cheeseman, MD
Bellenie, BR
van Montfort, RLM
Newton, GK
Burke, R
Collins, I
Hoelder, S
Mirza, A
McAndrew, PC
Thapaliya, A
Pierrat, OA
Stubbs, M
Hahner, T
Chessum, NEA
Innocenti, P
Caldwell, J
Cheeseman, MD
Bellenie, BR
van Montfort, RLM
Newton, GK
Burke, R
Collins, I
Hoelder, S
Document Type
Journal Article
Date
2023-08-10
Date Accepted
2023-07-19
Abstract
High hit rates from initial ligand-observed NMR screening can make it challenging to prioritize which hits to follow up, especially in cases where there are no available crystal structures of these hits bound to the target proteins or other strategies to provide affinity ranking. Here, we report a reproducible, accurate, and versatile quantitative ligand-observed NMR assay, which can determine Kd values of fragments in the affinity range of low μM to low mM using transverse relaxation rate R2 as the observable parameter. In this study, we examined the theory and proposed a mathematical formulation to obtain Kd values using non-linear regression analysis. We designed an assay format with automated sample preparation and simplified data analysis. Using tool compounds, we explored the assay reproducibility, accuracy, and detection limits. Finally, we used this assay to triage fragment hits, yielded from fragment screening against the CRBN/DDB1 complex.
Citation
Journal of Medicinal Chemistry, 2023, 66 (15), pp. 10617 - 10627
Source Title
Journal of Medicinal Chemistry
Publisher
AMER CHEMICAL SOC
ISSN
0022-2623
eISSN
1520-4804
1520-4804
1520-4804
Collections
Research Team
Hit Discov Struct Design
Medicinal Chemistry 4
Medicinal Chemistry 3
Medicinal Chemistry 4
Medicinal Chemistry 3