Fluorescence Polarization for the Evaluation of Small-Molecule Inhibitors of PCAF BRD/Tat-AcK50 Association

ChemMedChem ◽  
2014 ◽  
Vol 9 (5) ◽  
pp. 928-931 ◽  
Author(s):  
Ping Hu ◽  
Xinghui Wang ◽  
Baiqun Zhang ◽  
Shuai Zhang ◽  
Qiang Wang ◽  
...  
Keyword(s):  
Small Molecule ◽  
Fluorescence Polarization ◽  
Small Molecule Inhibitors

scholarly journals Optimization of Fluorescently Labeled Nrf2 Peptide Probes and the Development of a Fluorescence Polarization Assay for the Discovery of Inhibitors of Keap1-Nrf2 Interaction

2011 ◽  
Vol 17 (4) ◽  
pp. 435-447 ◽  
Author(s):  
Daigo Inoyama ◽  
Yu Chen ◽  
Xinyi Huang ◽  
Lesa J. Beamer ◽  
Ah-Ng Tony Kong ◽  
...  
Keyword(s):  
Small Molecule ◽  
Fluorescence Polarization ◽  
High Throughput Screening ◽  
Small Molecule Inhibitors ◽  
Dynamic Range ◽  
Antioxidant Response ◽  
Binding Affinities ◽  
Screening Assay ◽  
High Throughput Screening Assay ◽  
Peptide Amide

Activation of the antioxidant response element (ARE) upregulates enzymes involved in detoxification of electrophiles and reactive oxygen species. The induction of ARE genes is regulated by the interaction between redox sensor protein Keap1 and the transcription factor Nrf2. Fluorescently labeled Nrf2 peptides containing the ETGE motif were synthesized and optimized as tracers in the development of a fluorescence polarization (FP) assay to identify small-molecule inhibitors of the Keap1-Nrf2 interaction. The tracers were optimized to increase the dynamic range of the assay and their binding affinities to the Keap1 Kelch domain. The binding affinities of Nrf2 peptide inhibitors obtained in our FP assay using FITC-9mer Nrf2 peptide amide as the probe were in good agreement with those obtained previously by a surface plasmon resonance assay. The FP assay exhibits considerable tolerance toward DMSO and produced a Z′ factor greater than 0.6 in a 384-well format. Further optimization of the probe led to cyanine-labeled 9mer Nrf2 peptide amide, which can be used along with the FITC-9mer Nrf2 peptide amide in a high-throughput screening assay to discover small-molecule inhibitors of Keap1-Nrf2 interaction.

scholarly journals A Suite of Biochemical Assays for Screening RNA Methyltransferase BCDIN3D

2016 ◽  
Vol 22 (1) ◽  
pp. 32-39
Author(s):  
Levi L. Blazer ◽  
Fengling Li ◽  
Steven Kennedy ◽  
Yujun George Zheng ◽  
Cheryl H. Arrowsmith ◽  
...  
Keyword(s):  
Surface Plasmon Resonance ◽  
Surface Plasmon ◽  
Small Molecule ◽  
Plasmon Resonance ◽  
Fluorescence Polarization ◽  
Small Molecule Inhibitors ◽  
Biochemical Assays ◽  
Differential Scanning Fluorimetry ◽  
Small Molecule Modulators ◽  
Displacement Assays

BCDIN3D is an RNA-methyltransferase that O-methylates the 5′ phosphate of RNA and regulates microRNA maturation. To discover small-molecule inhibitors of BCDIN3D, a suite of biochemical assays was developed. A radiometric methyltransferase assay and fluorescence polarization–based S-adenosylmethionine and RNA displacement assays are described. In addition, differential scanning fluorimetry and surface plasmon resonance were used to characterize binding. These assays provide a comprehensive package for the development of small-molecule modulators of BCDIN3D activity.

A Fluorescence Polarization Based Screening Assay for Identification of Small Molecule Inhibitors of the PICK1 PDZ Domain

2011 ◽  
Vol 14 (7) ◽  
pp. 590-600 ◽  
Author(s):  
Thor S. Thorsen ◽  
Kenneth L. Madsen ◽  
Tino Dyhring ◽  
Anders Bach ◽  
Dan Peters ◽  
...  
Keyword(s):  
Small Molecule ◽  
Fluorescence Polarization ◽  
Small Molecule Inhibitors ◽  
Pdz Domain ◽  
Screening Assay
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