Jeremy Schofield
Site directed mutagenesis studies on HIV-1 reverse transcriptase (RT) enlighten the mechanism of action of a new Ribonuclease H/DNA polymerase RT dual inhibitor
ESPOSITO, FRANCESCA;CORONA, ANGELA;Rita Meleddu;DISTINTO, SIMONA;MACCIONI, ELIAS;
2014-01-01
Abstract
The principal target in the Human Immunodeficiency Virus therapy is the viral protein reverse transcriptase (RT). During the viral replication, RT accomplishes the DNA synthesis starting from the RNA viral genome by two associated functions: DNA polymerase and ribonuclease H (RNase H). Both activities are essential for viral replication and validated targets. At this time, none of the approved antiviral drugs acts on both activities, although few classes of compounds have been reported to inhibit both RT functions in vitro. In a previous virtual screening we identified a new isatine-based scaffold for dual functions RT inhibitors that was further derivatized. In particular, the isatine-derivative RMNC6 inhibited the RT-associated RNase H and DNA polymerase activities with IC50 values of 1.3 and 9.8 µM concentration, respectively. Blind docking analysis suggested that RMNC6 could bind to two different RT pockets. The first one is located close to the DNA polymerase catalytic site, contiguous to the NNRTI binding site. The second one is located in the RNase H domain, between the RNase H active site and the substrate-handle region close to the interface of the two subunits p66 and p51. Conserved residues close to the primer grip regions were individuated as potentially critical for the inhibitor binding and mutated. Mutagenesis results individuated few amino acid residues in the RNase H domain that are essential for RMNC6 inhibition of the RNase H function but do not influence its inhibition of the DNA polymerase function. In conclusion, we characterized the mechanism of action for the isatine-derivatived RNase H/RDDP dual inhibitor RMNC6, opening the way for further characterization of its binding pocket needed to allow further rational optimization of the scaffold.Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.