Abstract
Protein-Observed Fluorine NMR (PrOF NMR) spectroscopy is an emerging technique for screening and characterizing small-molecule–protein interactions. The choice of which amino acid to label for PrOF NMR can be critical for analysis. Here we report the first use of a protein containing two different fluoroaromatic amino acids for NMR studies. Using the KIX domain of the CBP/p300 as a model system, we examine ligand binding of several small-molecule compounds elaborated from our previous fragment screen and identify a new ligand binding site distinct from those used by native transcription factors. This site was further supported by computational modeling (FTMap and Schrödinger) and 1H,15N HSQC/HMQC NMR spectroscopy. Metabolic labeling with multiple fluorinated amino acids provides useful probes for further studying ligand binding and has led to new insight for allosterically regulating transcription-factor protein interactions with small-molecule ligands.
Original language | English (US) |
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Pages (from-to) | 963-969 |
Number of pages | 7 |
Journal | ChemBioChem |
Volume | 19 |
Issue number | 9 |
DOIs | |
State | Published - May 4 2018 |
Bibliographical note
Funding Information:This project was supported by the University of Minnesota, the NSF-CAREER Award CHE-1352091, the NIH training grant T32-GM08700, and the UMN Interdisciplinary Doctoral Fellowship (C.T.G.) We gratefully thank Prof. Sandor Vajda for helpful discussions on the applications of FTMap.
Publisher Copyright:
© 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Keywords
- NMR spectroscopy
- allosterism
- biomolecular NMR
- fluorine
- protein–protein interactions