Development and Validation of 2D Difference Intensity Analysis for Chemical Library Screening by Protein-Detected NMR.
 

Development and Validation of 2D Difference Intensity Analysis for Chemical Library Screening by Protein-Detected NMR.

http://www.bionmr.com//www.ncbi.nlm....x30_orange.png Related Articles Development and Validation of 2D Difference Intensity Analysis for Chemical Library Screening by Protein-Detected NMR.

Chembiochem. 2017 Dec 13;:

Authors: Egner JM, Jensen DR, Olp MD, Kennedy NW, Volkman BF, Peterson FC, Smith BC, Hill RB

Abstract
An academic chemical screening approach was developed and a 352-chemical fragment library screened against 3 different protein targets using 2D protein-detected NMR. The approach was optimized against 2 protein targets with known ligands, CXCL12 and BRD4. Principal component analysis reliably identified compounds that induced NMR crosspeak broadening, but did not unambiguously identify ligands with specific affinity (hits). For improved hit detection, a novel scoring metric - difference intensity analysis (DIA) -*was devised that sums all positive and negative intensities from 2D difference spectra. Applying DIA quickly discriminated potential ligands from compounds inducing NMR crosspeak broadening and other nonspecific effects. Subsequent NMR titrations validated chemotypes important for binding to CXCL12 and BRD4. A novel target, mitochondrial fission protein Fis1, was screened and 6 hits were identified using DIA. Screening these diverse protein targets identified quinones and catechols that induced NMR crosspeak broadening, hampered NMR analyses, but are currently not computationally identified as pan-assay interference compounds. The results establish a streamlined screening workflow that can easily be scaled and adapted as part of a larger screening pipeline to identify fragment hits and assess relative binding affinities in the range of 0.3 - 1.6 mM. DIA may prove useful in library screening and other applications where NMR chemical shift perturbations are measured.


PMID: 29239081 [PubMed - as supplied by publisher]



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