Back to Journals » Drug Design, Development and Therapy » Volume 7

Development of a robust cell-based high-throughput screening assay to identify targets of HIV-1 viral protein R dimerization

Authors Zych C, Domling A, Ayyavoo V

Received 18 February 2013

Accepted for publication 2 April 2013

Published 24 May 2013 Volume 2013:7 Pages 403—412

DOI https://doi.org/10.2147/DDDT.S44139

Checked for plagiarism Yes

Review by Single-blind

Peer reviewer comments 2


Courtney Zych,1 Alexander Domling,2 Velpandi Ayyavoo1

1Department of Infectious Diseases and Microbiology, Graduate School of Public Health, University of Pittsburgh, Pittsburgh, PA, USA; 2Department of Pharmacology, University of Pittsburgh, Pittsburgh, PA, USA

Abstract: Targeting protein–protein interactions (PPI) is an emerging field in drug discovery. Dimerization and PPI are essential properties of human immunodeficiency virus (HIV)-1 proteins, their mediated functions, and virus biology. Additionally, dimerization is required for the functional interaction of HIV-1 proteins with many host cellular components. In this study, a bimolecular fluorescence complementation (BiFC)-based screening assay was developed that can quantify changes in dimerization, using HIV-1 viral protein R (Vpr) dimerization as a "proof of concept." Results demonstrated that Venus Vpr (generated by BiFC Vpr constructs) could be competed off in a dose-dependent manner using untagged, full-length Vpr as a competitor molecule. The change in signal intensity was measured quantitatively through flow cytometry and fluorescence microscopy in a high content screening assay. High content imaging was used to screen a library of small molecules for an effect on Vpr dimerization. Among the tested molecules, a few of the small molecules demonstrate an effect on Vpr dimerization in a dose-dependent manner.

Keywords: BiFC, protein–protein interaction, HIV-1 Vpr, dimerization, drug targets

Creative Commons License This work is published and licensed by Dove Medical Press Limited. The full terms of this license are available at https://www.dovepress.com/terms.php and incorporate the Creative Commons Attribution - Non Commercial (unported, v3.0) License. By accessing the work you hereby accept the Terms. Non-commercial uses of the work are permitted without any further permission from Dove Medical Press Limited, provided the work is properly attributed. For permission for commercial use of this work, please see paragraphs 4.2 and 5 of our Terms.

Download Article [PDF]  View Full Text [HTML][Machine readable]