Fluorine Pseudocontact Shifts Used for Characterizing the Protein–Ligand Interaction Mode in the Limit of NMR Intermediate Exchange

• Published in: Angewandte Chemie International Edition 2017-10, Vol.56 (42), p.12982-12986
• Main Authors: Gao, Jia, Liang, E, Ma, Rongsheng, Li, Fudong, Liu, Yixiang, Liu, Jiuyang, Jiang, Ling, Li, Conggang, Dai, Haiming, Wu, Jihui, Su, Xuncheng, He, Wei, Ruan, Ke
• Format: Article
• Language: English
• Subjects: Binding Sites; bromodomains; Chelating Agents - chemistry; chemical exchange saturation transfer; Crystal structure; Exchanging; Fluorination; Fluorine; Fluorine - chemistry; Humans; Inhibitors; Lanthanoid Series Elements - chemistry; Ligands; Molecular Docking Simulation; Mutagenesis, Site-Directed; NMR; NMR spectroscopy; Nuclear magnetic resonance; Nuclear Magnetic Resonance, Biomolecular; Proteins; pseudocontact shifts; Saturation; Spatial data; Structural analysis; Transcription Factors - chemistry; Transcription Factors - genetics; Transcription Factors - metabolism
• ISSN: 1433-7851; 1521-3773
• DOI: 10.1002/anie.201707114

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 approach fills the gap of the NMR structural characterization of lead‐like inhibitors with moderate affinities to target proteins, which are essential for structure‐guided hit‐to‐lead evolution. The severe line broadening in the intermediate exchange limits the applicability of NMR spectroscopy for interrogating the interaction modes of lead‐like inhibitors with moderate affinities to target proteins. A 19F chemical exchange saturation transfer approach is used to retrieve the low‐populated bound‐state 19F pseudocontact shifts, which enable the identification of the best binding pose of the BRM bromodomain inhibitor.