Identification of a small-molecule binding site at the dimer interface of the HIV integrase catalytic domain

Abstract

Integration of the reverse‐transcribed HIV cDNA into the host DNA is a required step in viral replication. The virus‐encoded integrase protein catalyzes the initial DNA breaking and joining reactions that mediate cDNA integration. Here, the identification by X‐ray crystallography of a small‐molecule binding site on the integrase catalytic domain is reported. The small‐molecule family studied consists of a core of arsenic or phosphorus surrounded by four aromatic groups. Two arsenic derivatives were visualized bound to integrase. In each case, two molecules bound at symmetry‐related sites on the catalytic domain dimer interface. The first compound studied, tetraphenyl arsonium, did not inhibit integrase. However, a synthetic compound substituting a catechol for one of the phenyl rings, dihydroxyphenyltriphenylarsonium, bound to the same site and did inhibit the enzyme. Changes in the vicinity of the catalytic site were seen with the inhibitory compound only, potentially explaining its mechanism of action. Further substituting phosphonium for arsonium yielded a compound with an IC50 in the low micromolar range. These findings may be useful in designing new inhibitors of integrase, which is at present the only one of the three HIV enzymes for which clinically useful inhibitors are not available.

Identification of an inhibitor-binding site to HIV-1 integrase with affinity acetylation and mass spectrometry

“…Comparison of Acetylated-Inhibitor with 3,4-dihydroxyphenyltriphenylarsonium bromide, an HIV-1 IN inhibitor that has been shown to also bind at the IN dimer interface, is interesting (32). Our data indicate that Acetylated-Inhibitor binds deep into the dimer interface pocket owing to its ''slim'' structure (see Figs.…”

“…However, this structure did not include the cognate DNA substrate, which is believed to contribute to the inhibition through formation of a ternary inhibitor:protein:DNA complex. In separate work, a small molecule (3,4-dihydroxyphenyltriphenylarsonium bromide) inhibitor was shown to bind at the HIV-1 IN dimer interface (32). These structural analyses were performed by using the core domain of IN rather than the full-length protein (31,32).…”

“…In separate work, a small molecule (3,4-dihydroxyphenyltriphenylarsonium bromide) inhibitor was shown to bind at the HIV-1 IN dimer interface (32). These structural analyses were performed by using the core domain of IN rather than the full-length protein (31,32). Therefore, there is a need to devise new approaches that are complementary to crystallography and NMR to enable rapid and accurate mapping of inhibitor:protein contacts under biologically relevant conditions.…”

Integrase, LEDGF/p75 and HIV replication

“…In fact, prior to the recognition of this region as the p75 binding site, there was evidence it could contain hotspots for small molecule binding. Molteni et al [168] reported the structures of two related small molecules (tetraphenylarsonium and dihydroxyphenyltriphenylarsonium) bound to the IN CCD, where they were found to occupy what is now recognized as the IBD-binding cleft (compare panel b of Fig. 5 with Figure 3b of Molteni et al, which shows a similarly rotated view of the CCD).…”

Discovery of BI 224436, a Noncatalytic Site Integrase Inhibitor (NCINI) of HIV-1

“…Building on the work of Dyda et al, 12 we performed X-ray crystallographic experiments that demonstrated that this series of compounds bound to a hydrophobic pocket at the dimer interface of the CCD of IN (vide infra). This pocket was first identified as the binding site of tetraarylarsonium ions by X-ray crystallography 13 and later as a part of the recognition domain on IN for the lens epithelial derived growth factor (LEDGF/ p75), 14 a host cell protein involved in protection and trafficking of PICs to transcriptionally active regions of chromatin. 15 Since our series of inhibitors bound to a site remote from the active site and inhibited the 3′-processing activity of the enzyme, we have termed these molecules noncatalytic site integrase inhibitors (NCINIs).…”