Abstract

Styrylquinoline derivatives (SQ) efficiently inhibit the 3′-processing activity of integrase (IN) with IC₅₀ values of between 0.5 and 5 μM. We studied the mechanism of action of these compounds in vitro. First, we used steady-state fluorescence anisotropy to assay the effects of the SQ derivatives on the formation of IN-viral DNA complexes independently of the catalytic process. The IC₅₀ values obtained in activity and DNA-binding tests were similar, suggesting that the inhibition of 3′-processing can be fully explained by the prevention of IN-DNA recognition. SQ compounds act in a competitive manner, with K_i values of between 400 and 900 nM. In contrast, SQs did not inhibit 3′-processing when IN-DNA complexes were preassembled. Computational docking followed or not by molecular dynamics using the catalytic core of HIV-1 IN suggested a competitive inhibition mechanism, which is consistent with our previous data obtained with the corresponding Rous sarcoma virus domain. Second, we used preassembled IN-preprocessed DNA complexes to assay the potency of SQs against the strand transfer reaction, independently of 3′-processing. Inhibition occurred even if the efficiency was decreased by about 5- to 10-fold. Our results suggest that two inhibitor-binding modes exist: the first one prevents the binding of the viral DNA and then the two subsequent reactions (i.e., 3′-processing and strand transfer), whereas the second one prevents the binding of target DNA, thus inhibiting strand transfer. SQ derivatives have a higher affinity for the first site, in contrast to that observed for the diketo acids, which preferentially bind to the second one.