title: Structural and inhibition studies of the RNase H function of xenotropic murine leukemia virus-related virus reverse transcriptase.
abstract: RNase H inhibitors (RNHIs) have gained attention as potential HIV-1 therapeutics. Although several RNHIs have been studied in the context of HIV-1 reverse transcriptase (RT) RNase H, there is no information on inhibitors that might affect the RNase H activity of other RTs. We performed biochemical, virological, crystallographic, and molecular modeling studies to compare the RNase H function and inhibition profiles of the gammaretroviral xenotropic murine leukemia virus-related virus (XMRV) and Moloney murine leukemia virus (MoMLV) RTs to those of HIV-1 RT. The RNase H activity of XMRV RT is significantly lower than that of HIV-1 RT and comparable to that of MoMLV RT. XMRV and MoMLV, but not HIV-1 RT, had optimal RNase H activities in the presence of Mn²⁺ and not Mg²⁺. Using hydroxyl-radical footprinting assays, we demonstrated that the distance between the polymerase and RNase H domains in the MoMLV and XMRV RTs is longer than that in the HIV-1 RT by ∼3.4 Å. We identified one naphthyridinone and one hydroxyisoquinolinedione as potent inhibitors of HIV-1 and XMRV RT RNases H with 50% inhibitory concentrations ranging from ∼0.8 to 0.02 μM. Two acylhydrazones effective against HIV-1 RT RNase H were less potent against the XMRV enzyme. We also solved the crystal structure of an XMRV RNase H fragment at high resolution (1.5 Å) and determined the molecular details of the XMRV RNase H active site, thus providing a framework that would be useful for the design of antivirals that target RNase H.
authors.affiliation: Christopher S. Bond Life Sciences Center, University of Missouri, Columbia, Missouri, USA.
authors.forename: Karen A
authors.initials: KA
authors.lastname: Kirby
authors.forename: Bruno
authors.initials: B
authors.lastname: Marchand
authors.forename: Yee Tsuey
authors.initials: YT
authors.lastname: Ong
authors.forename: Tanyaradzwa P
authors.initials: TP
authors.lastname: Ndongwe
authors.forename: Atsuko
authors.initials: A
authors.lastname: Hachiya
authors.forename: Eleftherios
authors.initials: E
authors.lastname: Michailidis
authors.forename: Maxwell D
authors.initials: MD
authors.lastname: Leslie
authors.forename: Daniel V
authors.initials: DV
authors.lastname: Sietsema
authors.forename: Tracy L
authors.initials: TL
authors.lastname: Fetterly
authors.forename: Christopher A
authors.initials: CA
authors.lastname: Dorst
authors.forename: Kamalendra
authors.initials: K
authors.lastname: Singh
authors.forename: Zhengqiang
authors.initials: Z
authors.lastname: Wang
authors.forename: Michael A
authors.initials: MA
authors.lastname: Parniak
authors.forename: Stefan G
authors.initials: SG
authors.lastname: Sarafianos
mesh_terms: D000595:Amino Acid Sequence; D019380:Anti-HIV Agents / Q000494:pharmacology*; D002470:Cell Survival / Q000187:drug effects; D018360:Crystallography, X-Ray; D018983:DNA Footprinting; D054303:HIV Reverse Transcriptase / Q000037:antagonists & inhibitors; D006835:Hydrazones / Q000138:chemical synthesis / Q000494:pharmacology; D007202:Indicators and Reagents; D007546:Isoquinolines / Q000138:chemical synthesis / Q000494:pharmacology; D008274:Magnesium / Q000494:pharmacology; D008345:Manganese / Q000494:pharmacology; D008826:Microbial Sensitivity Tests; D008958:Models, Molecular; D008969:Molecular Sequence Data; D008979:Moloney murine leukemia virus / Q000187:drug effects / Q000201:enzymology; D009287:Naphthyridines / Q000138:chemical synthesis / Q000494:pharmacology; D010957:Plasmids / Q000235:genetics; D012194:RNA-Directed DNA Polymerase / Q000737:chemistry*; D018894:Reverse Transcriptase Inhibitors / Q000494:pharmacology*; D016914:Ribonuclease H / Q000037:antagonists & inhibitors* / Q000737:chemistry* / Q000502:physiology*; D058008:Xenotropic murine leukemia virus-related virus / Q000201:enzymology*
chemical_list: D019380:Anti-HIV Agents; D006835:Hydrazones; D007202:Indicators and Reagents; D007546:Isoquinolines; D009287:Naphthyridines; D018894:Reverse Transcriptase Inhibitors; D008345:Manganese; D054303:HIV Reverse Transcriptase; D012194:RNA-Directed DNA Polymerase; D016914:Ribonuclease H; D008274:Magnesium