Inhibition of Foamy Virus Reverse Transcriptase by Human Immunodeficiency Virus Type 1 RNase H Inhibitors

Abstract

bRNase H plays an essential role in the replication of human immunodeficiency virus type 1 (HIV-1). Therefore, it is a promising target for drug development. However, the identification of HIV-1 RNase H inhibitors (RHIs) has been hampered by the open morphology of its active site, the limited number of available RNase H crystal structures in complex with inhibitors, and the fact that, due to the high concentrations of Mg 2؉ needed for protein stability, HIV-1 RNase H is not suitable for nuclear magnetic resonance (NMR) inhibitor studies. We recently showed that the RNase H domains of HIV-1 and prototype foamy virus (PFV) reverse transcriptases (RTs) exhibit a high degree of structural similarity. Thus, we examined whether PFV RNase H can serve as an HIV-1 RNase H model for inhibitor interaction studies. Five HIV-1 RHIs inhibited PFV RNase H activity at low-micromolar concentrations similar to those of HIV-1 RNase H, suggesting pocket similarity of the RNase H domains. NMR titration experiments with the PFV RNase H domain and the RHI RDS1643 (6-[1-(4-fluorophenyl)methyl-1H-pyrrol-2-yl)]-2,4-dioxo-5-hexenoic acid ethyl ester) were performed to determine its binding site. Based on these results and previous data, in silico docking analysis showed a putative RDS1643 binding region that reaches into the PFV RNase H active site. Structural overlays were performed with HIV-1 and PFV RNase H to propose the RDS1643 binding site in HIV-1 RNase H. Our results suggest that this approach can be used to establish PFV RNase H as a model system for HIV-1 RNase H in order to identify putative inhibitor binding sites in HIV-1 RNase H.

Identification of Highly Conserved Residues Involved in Inhibition of HIV-1 RNase H Function by Diketo Acid Derivatives

“…Molecular modeling studies on the RT RNase H domain predicted that our DKAs would interact with a number of residues surrounding the RNase H active site, i.e., R448, N474, Q475, Y501, and R557, most of which are highly conserved (45) as part of the RNase H primer grip motif (41,42,46). This is a singular pattern that is interesting for the large number of residues involved and is in agreement with recent observations on the interaction of RDS1643 and the PFV domain of RNase H (32). In fact, all of the crystal structures of RT/RNase H prototypes solved in complexes with active site RHIs show the ligands coordinating the metal cofactors (25,47) while exhibiting an orientation of binding that does not allow extended secondary interactions with amino acid side chains.…”

“…Recently, a new series of pyrrolyl DKAs that are active against both HIV-1 IN and RNase H has been reported (31), with acidic compounds being more effective with IN and ester counterparts being active with both enzymes, with no particular difference. Moreover, a recent study on RHI effects on and binding to prototype foamy virus (PFV) RT showed a putative RDS1643 binding region in the PFV RNase H active site and suggested, given the high level of structural similarity between PFV and HIV-1 RNase H domains, the possibility that RDS1643 could show similar interactions with the HIV-1 RNase H domain (32).…”

Synthetic α-Hydroxytropolones Inhibit Replication of Wild-Type and Acyclovir-Resistant Herpes Simplex Viruses

“…␤-Thujaplicinol had been tested against HSV because the structure and/or function of several enzymes involved in HSV DNA replication is consistent with that of NTS enzymes. Other viral enzymes in the nucleotidyltransferase superfamily include the HBV and HIV RNase H enzymes (31,32), and ␤-thujaplicinol has efficacy against HIV RNase H (37-41), HBV RNase H (42), and foamy virus (62). Here, 27 additional tropolones were screened against HSV-1 and HSV-2.…”

Hydroxylated Tropolones Inhibit Hepatitis B Virus Replication by Blocking Viral Ribonuclease H Activity

“…Previously, we identified the ␣-hydroxytropolone natural product ␤-thujaplicinol (compound 46) as an inhibitor of HBV RNaseH (14) based on its efficacy against HIV RNaseH (21)(22)(23)(24)(25). Here, 50 additional troponoids, including 36 ␣-hydroxytropolones, were screened against HBV RNaseH based on our previous results and the ability of this compound class to inhibit the HIV enzyme and to cross-inhibit other viruses such as the herpes simplex viruses (26) and foamy virus (39). Our goal was to determine whether tropolones are candidates for development as anti-HBV drugs and, if so, to derive initial structure-activity relationships (SAR) that could help guide their chemical optimization.…”