doi:10.1016/j.bmcl.2006.02.031 
Copyright © 2006 Elsevier Ltd All rights reserved.
A fast and robust 19F NMR-based method for finding new HIV-1 protease inhibitors
Silvia Frutosa, Teresa Tarragoa and Ernest Giralta, b, 
, 
aInstitut de Recerca Biomèdica (IRB-PCB), Parc Científic de Barcelona, Josep Samitier, 1-5. E-08028 Barcelona, Spain
bDepartament de Química Orgànica, Universitat de Barcelona, Martí i Franquès, 1. E-08028 Barcelona, Spain
Received 5 December 2005;
revised 8 February 2006;
accepted 9 February 2006.
Available online 6 March 2006.
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Abstract
The human immunodeficiency virus (HIV) which encodes, among other indispensable enzymes, an aspartic protease that is essential for viral maturation and replication. Numerous inhibitors of the protease have been developed. However, the eventual resistance of HIV-1 to these drugs implies a continuous battle to develop new inhibitors. Proposed herein is a robust, fast, and reliable method employing 19F NMR for the evaluation of the inhibitory activity of new compounds against HIV-1 protease.
Keywords: 19F NMR; HIV-1 protease; NMR screening; Protease inhibitors; Traditional Chinese Medicine
Scheme 1. (a) Enzymatic reaction of HIV-1 protease in the presence of a fluorinated substrate; (b) a model 19F NMR spectrum corresponding to partial enzymatic reaction.
Figure 1. Substrate 1 and 19F NMR spectra of its enzymatic reaction. (a) Fluorinated peptidyl substrate 1. (b) 19F NMR spectrum recorded for the substrate 1 after incubation with purified HIV-1 protease.
Figure 2. Substrate 2 and 19F NMR spectra of its enzymatic reaction. (a) Fluorinated peptidyl substrate 2. (b) 19F NMR spectra recorded for the substrate 2 after incubation with purified HIV-1 protease.
Figure 3. HPLC–MS chromatogram of the partial hydrolysis of substrate 2 by HIV-1 protease. Inset: ES+/MS spectra for the uncleaved peptide (top) and the product (bottom).
Figure 4. Synthetic inhibitors of HIV-1 protease. (a) Active site inhibitor, inhibitor 3. (b) Dimerization inhibitor, inhibitor 4.
Figure 5. 19F NMR spectra corresponding to the substrate 2 (a) in the absence of inhibitor; (b) in the presence of dimerization inhibitor, inhibitor 4; (c) in the presence of active site inhibitor, inhibitor 3.
Figure 6. 19F NMR spectra corresponding to the substrate 2 after incubation with purified HIV-1 protease for (a) 15 min in the absence of inhibitor; (b) 15 min in the presence of inhibitor 3 (c) 30 min in the absence of inhibitor; (d) 30 min in the presence of inhibitor 3; (e) 1 h in the absence of inhibitor (f) 1 h in the presence of inhibitor 3.
Figure 7. 19F NMR spectra corresponding to the substrate 2 (a) in the presence of Traditional Chinese Medicine extract 10 (b) in the presence of Traditional Chinese Medicine extract 16.
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