Compounds that interact with DNA or RNA generally act as inhibitors of enzymes that unwind DNA or RNA. In the present study we describe the synthesis and properties of some nucleoside analogues that interact with double-stranded DNA but that, in contrast, facilitate the unwinding reaction mediated by West Nile (
In the presented study the ribavirin-TP--an established inhibitor of the NTPase activity of the superfamily NTPase/helicases II--was investigated as an inhibitor of the unwinding activity of the hepatitis C virus (HCV) NTPase/helicase. The kinetics of the reaction revealed that ribavirin-TP reduces the turnover number of the helicase reaction by a mechanism that does not correspond to that of the inhibition of the NTPase activity. Our results suggest that derivatives of ribavirin-TP with enhanced stability towards hydrolytic attack may be effective inhibitors of the enzyme.
Hepatitis C virus (HCV), West Nile virus (WNV), Japanese encephalitis virus ( JEV), and dengue virus (DENV) are members of Flaviviridae. They are deadly human pathogens and new and better therapeutic strategies are desperately needed. However, there are considerable barriers to the development of anti-Flaviviridae therapeutics, including the persistence of the virus, its genetic diversity during replication in the host and, for example, in the case of HCV, the lack of reproducible infectious culture systems. Similarities in the key Flaviviridae enzymes (with respect to conserved subdomain organization and arrangement of structural motifs) may indicate the potential usefulness of ribonucleoside analogues in the treatment of these viral infections. Over the last few years our understanding of the Flaviviridae replication cycle and the key enzymes involved in it has increased, giving hope for development of new antiviral agents.The Flaviviridae genome is a positive-stranded RNA, with a single open reading frame that translates into a single polyprotein. This polyprotein is co-and post-translationally processed by cellular and viral proteases into structural and non-structural (NS) proteins (Rosenberg, 2001). Among the NS proteins, NS3 and NS5 -the major components of the so called 'replication complex' -appear to be the most promising targets for antiviral therapy because of their well-characterized enzymatic activities (Tan et al.,
In this report, we demonstrate the interaction of the non-structural protein 3 (NS3) of hepatitis C virus (HCV) with alkaloide tropolone (2-hydroxy-2,4,6-heptatriene-1-one) and its derivatives. The compounds were biochemically screened separately against the ATPase and helicase activities of HCV NS3. In the investigations presented, alkaloide tropolone and its derivatives significantly inhibited the helicase activity of the viral protein when using a DNA substrate, with 50% inhibitory concentration values within a low micromolar range. The results using the RNA substrate were unexpected -none of the tropolone derivatives excerted any modulating influence towards the unwinding activity. Surprisingly, no influence of the nucleoside triphosphatase (NTPase) turnover was observed. Evidence is presented confirming that these compounds do not act by blocking the NTP-binding site, but by occupying an additional allosteric regulatory site. Further mechanisms of action, particularly of some of the derivatives, are discussed.
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