Fragment screening using capillary electrophoresis (CEfrag) for hit identification of heat shock protein 90 ATPase inhibitors

Journal of Biomolecular Screening
Carol AustinDallas E Hughes

Abstract

CEfrag is a new fragment screening technology based on affinity capillary electrophoresis (ACE). Here we report on the development of a mobility shift competition assay using full-length human heat shock protein 90α (Hsp90α), radicicol as the competitor probe ligand, and successful screening of the Selcia fragment library. The CEfrag assay was able to detect weaker affinity (IC(50) >500 µM) fragments than were detected by a fluorescence polarization competition assay using FITC-labeled geldanamycin. The binding site of selected fragments was determined by co-crystallization with recombinant Hsp90α N-terminal domain and X-ray analysis. The results of this study confirm that CEfrag is a sensitive microscale technique enabling detection of fragments binding to the biological target in near-physiological solution.

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Citations

Oct 15, 2014·Expert Opinion on Drug Discovery·Gengyang YuanGraham B Jones
Dec 8, 2015·Analytical Chemistry·Rachel K HarstadMichael T Bowser
Jan 24, 2014·Drug Discovery Today. Technologies·Laura SilvianGnanasambandam Kumaravel
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Aug 28, 2020·Frontiers in Molecular Biosciences·Qingxin Li
Oct 30, 2020·Progress in Biophysics and Molecular Biology·David M WilsonDebanu Das

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Methods Mentioned

BETA
X-ray
electrophoresis
Assay
nuclear magnetic resonance
gel filtration
NMR

Software Mentioned

COOT
REFAMAC5
Libcheck
MolProbity
PHASER
CCP4
CEfrag
PROTIUM2

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