Duck hepatitis B virus: DNA polymerase and reverse transcriptase activities of replicative complexes isolated from liver and their inhibition in vitro.
The duck hepatitis B virus (DHBV)-associated activities of reverse transcriptase and DNA polymerase and their inhibition in vitro were studied. Replicative complexes (RCs) were isolated from DHBV-infected liver by gel chromatography followed by sucrose gradient centrifugation. The RCs were detected by dot blot hybridization, using radiolabeled cloned DHBV DNA as a probe, and by the incorporation of 32P-TTP in the presence of dATP, dCTP, dGTP, and Mg2+ (endogenous DNA polymerase activity). The endogenous DNA polymerase activity associated with RCs was further studied using exogenous templates: reverse transcriptase and DNA polymerase activities were demonstrated using as substrates 32P-TTP and poly(rA) p(dT)12 or poly(dA) p(dT)12-18, respectively. Both activities were biochemically characterized. Their inhibition by various antiviral agents was studied in vitro: actinomycin D, ara-ATP, aphidicolin, suramin, chloroquin, and phosphonoformate. Among these, suramin, chloroquin, phosphonoformate, and ara-ATP were shown to be potent inhibitors of viral reverse transcriptase and DNA polymerase. Studies are now in progress to establish their antiviral activity in vivo.
D000818:Animals; D004259:DNA-Directed DNA Polymerase / Q000302:isolation & purification*; D004372:Ducks; D006515:Hepatitis B virus / Q000201:enzymology* / Q000502:physiology; D006524:Hepatitis, Viral, Animal / Q000378:metabolism; D008099:Liver / Q000032:analysis; D019384:Nucleic Acid Synthesis Inhibitors; D012194:RNA-Directed DNA Polymerase / Q000302:isolation & purification*; D018894:Reverse Transcriptase Inhibitors; D014764:Viral Proteins / Q000302:isolation & purification*; D014779:Virus Replication
D019384:Nucleic Acid Synthesis Inhibitors; D018894:Reverse Transcriptase Inhibitors; D014764:Viral Proteins; D012194:RNA-Directed DNA Polymerase; D004259:DNA-Directed DNA Polymerase