Dual Labeling of the CBP/p300 KIX Domain for 19F NMR Leads to Identification of a New Small-Molecule Binding Site

Clifford T. Gee, Keith E. Arntson, Edward J. Koleski, Rachel Lynn Staebell, William C Pomerantz

Research output: Contribution to journalArticlepeer-review

5 Scopus citations

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 languageEnglish (US)
Pages (from-to)963-969
Number of pages7
JournalChemBioChem
Volume19
Issue number9
DOIs
StatePublished - 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.

Keywords

  • NMR spectroscopy
  • allosterism
  • biomolecular NMR
  • fluorine
  • protein–protein interactions

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