Interfacial peptide inhibitors of HIV-1 integrase activity and dimerization

Zhao, Lei; O’Reilly, Mary K.; Shultz, Michael D.; Chmielewski, Jean

doi:10.1016/s0960-894x(03)00040-4

Cited by 57 publications

(49 citation statements)

References 19 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…13 Proof of principle for this inhibitory strategy has previously been reported, generally through the use of peptides. [13][14][15][16][20][21][22] In our workflow, we included a GST-His 6 AlphaScreen counterscreen to rapidly and confidently purge initial hit lists of false positives.…”

Section: Discussionmentioning

confidence: 99%

Development of an AlphaScreen-Based HIV-1 Integrase Dimerization Assay for Discovery of Novel Allosteric Inhibitors

et al. 2012

View full text Add to dashboard Cite

In recent years, HIV-1 integrase (IN) has become an established target in the field of antiretroviral drug discovery. However, its sole clinically approved inhibitor, the integrase strand transfer inhibitor (INSTI) raltegravir, has a surprisingly low genetic barrier for resistance. Furthermore, the only two other integrase inhibitors currently in advanced clinical trials, elvitegravir and dolutegravir, share its mechanism of action and certain resistance pathways. To maintain a range of treatment options, drug discovery efforts are now turning toward allosteric IN inhibitors, which should be devoid of cross-resistance with INSTIs. As IN requires a precise and dynamic equilibrium between several oligomeric species for its activities, the modulation of this equilibrium presents an interesting allosteric target. We report on the development, characterization, and validation of an AlphaScreen-based assay for high-throughput screening for modulators of HIV-1 IN dimerization. Compounds identified as hits in this assay proved to act as allosteric IN inhibitors. Additionally, the assay offers a flexible platform to study IN dimerization.

show abstract

Section: Discussionmentioning

confidence: 99%

Development of an AlphaScreen-Based HIV-1 Integrase Dimerization Assay for Discovery of Novel Allosteric Inhibitors

et al. 2012

View full text Add to dashboard Cite

show abstract

“…The reaction mixtures were treated with the cross-linking agent BS3 (25, 50, or 100 eq, Pierce) for 30 min at 37°C. The reaction mixtures were denatured and analyzed by 10% SDS-PAGE (37,38).…”

Section: Methodsmentioning

confidence: 99%

The Structure and Interactions of the Proline-rich Domain of ASPP2

Rotem

Katz

Benyamini

et al. 2008

Journal of Biological Chemistry

View full text Add to dashboard Cite

ASPP2 is a pro-apoptotic protein that stimulates the p53-mediated apoptotic response. The C terminus of ASPP2 contains ankyrin (Ank) repeats and a SH3 domain, which mediate its interactions with numerous partner proteins such as p53, NFB, and Bcl-2. It also contains a proline-rich domain (ASPP2 Pro), whose structure and function are unclear. Here we used biophysical and biochemical methods to study the structure and the interactions of ASPP2 Pro, to gain insight into its biological role. We show, using biophysical and computational methods, that the ASPP2 Pro domain is natively unfolded. We found that the ASPP2 Pro domain interacts with the ASPP2 Ank-SH3 domains, and mapped the interaction sites in both domains. Using a combination of peptide array screening, biophysical and biochemical techniques, we found that ASPP2 Ank-SH3, but not ASPP2 Pro, mediates interactions of ASPP2 with peptides derived from its partner proteins. ASPP2 Pro-Ank-SH3 bound a peptide derived from its partner protein NFB weaker than ASPP2 Ank-SH3 bound this peptide. This suggested that the presence of the proline-rich domain inhibited the interactions mediated by the Ank-SH3 domains. Furthermore, a peptide from ASPP2 Pro competed with a peptide derived from NFB on binding to ASPP2 Ank-SH3. Based on our results, we propose a model in which the interaction between the ASPP2 domains regulates the intermolecular interactions of ASPP2 with its partner proteins.

show abstract

“…In 12 of the studies published in 2003 using recombinant retroviral integrases, four were performed with Mg 2ϩ as a divalent metal cofactor (Cherepanov et al, 2003;Singh et al, 2003a,b;Zhuang et al, 2003), whereas five studies used Mn 2ϩ (Bao et al, 2003;Priet et al, 2003;Zagarian et al, 2003;Zhang et al, 2003;Zhao et al, 2003). Three other studies were performed with Mg 2ϩ as well as Mn 2ϩ (Bischerour et al, 2003;King et al, 2003;Podtelezhnikov et al, 2003).…”

mentioning

confidence: 99%

Metal-Dependent Inhibition of HIV-1 Integrase by β-Diketo Acids and Resistance of the Soluble Double-Mutant (F185K/C280S)

Marchand

Johnson²,

Karki³

et al. 2003

Mol Pharmacol

125

106

View full text Add to dashboard Cite

The ␤-diketo acids (DKAs) represent a major advance for anti-HIV-1 integrase drug development. We compared the inhibition of HIV-1 integrase by six DKA derivatives using the wild-type enzyme or the double-mutant F185K/C280S, which has been previously used for crystal structure determinations. With the wild-type enzyme, we found that DKAs could be classified into two groups: those similarly potent in the presence of magnesium and manganese and those potent in manganese and relatively ineffective in the presence of magnesium. Both the aromatic and the carboxylic or tetrazole functions of DKAs determined their metal selectivity. The F185K/C280S enzyme was markedly more active in the presence of manganese than magnesium. The F185K/C280S integrase was also relatively resistant to the same group of DKAs that were potent in the presence of magnesium with the wild-type enzyme. Resistance was caused by a synergistic effect from both the F185K and C280S mutations. Molecular modeling and docking suggested metal-dependent differences for binding of DKAs. Molecular modeling also indicated that the tetrazole or the azido groups of some derivatives could directly chelate magnesium or manganese in the integrase catalytic site. Together, these experiments suggest that DKAs recognize conformational differences between wild-type and the double-mutant HIV-1 integrase, because they chelate the magnesium or manganese in the enzyme active site and compete for DNA binding.

show abstract

Interfacial peptide inhibitors of HIV-1 integrase activity and dimerization

Cited by 57 publications

References 19 publications

Development of an AlphaScreen-Based HIV-1 Integrase Dimerization Assay for Discovery of Novel Allosteric Inhibitors

Development of an AlphaScreen-Based HIV-1 Integrase Dimerization Assay for Discovery of Novel Allosteric Inhibitors

The Structure and Interactions of the Proline-rich Domain of ASPP2

Metal-Dependent Inhibition of HIV-1 Integrase by β-Diketo Acids and Resistance of the Soluble Double-Mutant (F185K/C280S)

Contact Info

Product

Resources

About