Use of Fluorine Pseudocontact Shifts to Characterize the Protein-Ligand Interaction Mode in the Limit of NMR Intermediate Exchange
 

Use of Fluorine Pseudocontact Shifts to Characterize the Protein-Ligand Interaction Mode in the Limit of NMR Intermediate Exchange


The characterization of protein-ligand interaction modes becomes recalcitrant in the NMR intermediate exchange regime as the interface resonances are broadened beyond detection. Here, we determined the 19F low-populated bound-state pseudocontact shifts (PCSs) of mono- and di-fluorinated inhibitors of the BRM bromodomain using a highly-skewed protein/ligand ratio. The bound-state 19F PCSs were retrieved from 19F chemical exchange saturation transfer (CEST) in the presence of the lanthanide-labeled protein, which was termed the 19F PCS-CEST approach. These PCSs enriched in spatial information enabled the identification of best-fitting poses, which agree well with the crystal structure of a more soluble analog in complex with the BRM bromodomain. This 19F PCS-CEST approach fills the gap of the NMR structural characterization of lead-like inhibitors with intermediate affinities, which are essential for structure-guided hit-to-lead evolution.

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