scholarly journals Small Dumbbell Oligonucleotides Inhibitors of RNase H Activity of MOMULV Reverse Transcriptase

2011 ◽  
Vol 8 (2) ◽  
pp. 629-634
Author(s):  
Ajay Kumar
Keyword(s):  
Reverse Transcriptase ◽  
Murine Leukemia Virus ◽  
Dissociation Constants ◽  
Leukemia Virus ◽  
Rnase H ◽  
Moloney Murine Leukemia Virus ◽  
Mobility Shift ◽  
Hiv 1 ◽  
Murine Leukemia

The small dumbbell oligonucleotides containing loops of phosphodiester (OL-1), two trimethylene, C3moieties in each loop (OL-2) and phosphorothioate (OL-3) linkages were synthesized. Incubation of OL-1 and OL-2 with S-1 nuclease generated break down products whereas incubation of OL-3 did not result in significant cleavage. Their binding to moloney murine leukemia virus reverse transcriptase was evaluated by PAGE band mobility shift assays. The OL-3 bound more strongly to the reverse transcriptase than OL-1 and OL-2. The dissociation constants evaluated using PAGE band mobility shift assays were of the order of 10-7. Investigation of inhibition of RNase H activity of reverse transcriptase showed that the OL-3 is a better inhibitor of the retroviral RNase H activity than both OL-1 and OL-2. Thus OL-3 may be used as RNase H inhibitor. Our studies demonstrated that this particularly designed oligonucleotide (OL-3) displays an IC50of 25 nM in its inhibition on the reverse transcriptase RNase H activity, a magnitude lower than that of first nucleotide reverse transcriptase of HIV-1, tenofovir, introduced by Gilead Science in the market.

scholarly journals Binding of Dumbbell Oligonucleotides to MoMuLV Reverse Transcriptase: Inhibitory Properties of RNase H Activity

2010 ◽  
Vol 7 (3) ◽  
pp. 701-708
Author(s):  
Ajay Kumar
Keyword(s):  
Reverse Transcriptase ◽  
Dissociation Constants ◽  
Leukemia Virus ◽  
Rnase H ◽  
Moloney Murine Leukemia Virus ◽  
Mobility Shift ◽  
S1 Nuclease ◽  
Similar Affinity ◽  
Hiv 1 ◽  
Murine Leukemia

Dumbbell oligonucleotides with loops of various chemistry were synthesized. Incubation of dumbbell oligonucleotides containing phosphorothioate bonds or trimethylene phosphate linkages in loops with S1 nuclease did not result in significant cleavage under conditions which led to the degradation of dumbbell oligonucleotide containing phophodiester bonds in the loops. The binding of reverse transcriptase of Moloney Murine Leukemia Virus (MoMuLV) was evaluated with all the five oligonucleotides. The protein binds to all the dumbbell oligonucleotides with similar affinity. The dissociation constants evaluated using PAGE band mobility shift assays were of the order of 10-7. The inhibitory properties of the retroviral RNase H activity was evaluated using3H –UTP-labeled RNA:RNA-DNA hybrid. It was found that the best dumbbell oligonucleotide, inhibitor contained phosphorothioate residues in both the loops. Our value studies demonstrated that this particularly designed oligonucleotide displays an IC50of 18 nM in its inhibition on the reverse transcriptase RNase H activity, a magnitude lower than that of first nucleotide reverse transcriptase of HIV-1, tenofovir, introduced by Gilead Science in the market.

scholarly journals Structural and Inhibition Studies of the RNase H Function of Xenotropic Murine Leukemia Virus-Related Virus Reverse Transcriptase

2012 ◽  
Vol 56 (4) ◽  
pp. 2048-2061 ◽  
Author(s):  
Karen A. Kirby ◽  
Bruno Marchand ◽  
Yee Tsuey Ong ◽  
Tanyaradzwa P. Ndongwe ◽  
Atsuko Hachiya ◽  
...  
Keyword(s):  
Reverse Transcriptase ◽  
Murine Leukemia Virus ◽  
Leukemia Virus ◽  
Rnase H ◽  
Moloney Murine Leukemia Virus ◽  
Xenotropic Murine Leukemia ◽  
Related Virus ◽  
Hydroxyl Radical Footprinting ◽  
Hiv 1 ◽  
Murine Leukemia

ABSTRACTRNase 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.

scholarly journals Preparation and characterization of the RNase H domain of Moloney murine leukemia virus reverse transcriptase

2015 ◽  
Vol 113 ◽  
pp. 44-50 ◽  
Author(s):  
Kosaku Nishimura ◽  
Kanta Yokokawa ◽  
Tetsuro Hisayoshi ◽  
Kosuke Fukatsu ◽  
Ikumi Kuze ◽  
...  
Keyword(s):  
Reverse Transcriptase ◽  
Murine Leukemia Virus ◽  
Leukemia Virus ◽  
Rnase H ◽  
Moloney Murine Leukemia Virus ◽  
Murine Leukemia
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