19F NMR for Fragment-Based Small Molecule Discovery

Fragment screening by 19F NMR

Fluorinated Proteins for Transcription Factor Ligand Discovery Using Protein-observed Fluorine (PrOF) NMR Spectroscopy.

      To begin a medicinal chemistry campaign for PPIs, a significant challenge is to first discover lead molecules. NMR screening has become a highly valued method due to its ability to quantify a wide range of affinities for small molecules, particularly weak ligands in the area of fragment-based drug discovery (FBDD). We have demonstrated a new protein-observed fluorine NMR method (PrOF NMR) for screening and characterizing ligand binding at PPI sites.   We replace aromatic amino acids due to their enrichment at PPI interfaces, and employ singly-labeled fluorinated residues to minimize perturbation to the native recognition events. This strategy readily yields 1D-NMR spectra that report directly on protein structure from small to medium-sized proteins and is ideal for FBDD.

    We have applied our approach to understand protein-protein interactions and allosteric communication in the kinase inducible interaction domain (KIX) of coactivator CBP. We have demonstrated the use of PrOF NMR for ligand discovery and have recently characterized a new binding site on KIX distinct from natural transcription factors.  We currently are exploring the biological significance of this new site for regulating transcription through development of improved inhibitors, binding site analysis, and investigation of the KIX allosteric network through the lens of aromatic side chains.  Our small molecule studies complement our efforts using macrocyclic peptides as new ligands for the KIX domain in Project Area 3.  Finally, using KIX as a model system, we are developing our PrOF NMR approach for enabling faster data collection, testing new fluorinated amino acids, and quantifying protein dynamics.  

For Further Reading see:

4. “Dual Labeling of the CBP/p300 KIX domain for 19F NMR leads to identification of a new small molecule binding site.” C. T. Gee, K. E. Arntson, E. J. Koleski, Rachel Staebell, W. C. K. Pomerantz*, ChemBioChem 2018, 19, 963-9.

3. Protein-Observed 19F NMR for fragment screening, affinity quantification, and druggability assessment.” C. T. Gee, K. E. Arntson, L. M. Hawk, A. Wisniewski, A. K. Urick, N. K. Mishra, W. C. K. Pomerantz*, Nat. Protoc. 2016, 11, 1414-27

2. Fragment Screening and Druggability Assessment for the CBP/p300 KIX Domain Via Protein Observed 19F NMR” C. T. Gee, E. J. Koleski W. C. Pomerantz*.  Angewandte Chemie Int Ed. 2015 54. 3735-9

1. “Profiling the Dynamic Interfaces of Fluorinated Transcription Complexes for Ligand Discovery and Characterization” W.C. Pomerantz, N. Wang; R. Wang, A. K. Lipinksi, T. Cierpicki, A. K. Mapp. ACS. Chem. Biol., 2012, 7, 1345-50.