RNA helicase activity of NS3 proteins of hepatitis C virus and hepatitis G virus

Abstract

The Hepatitis C virus (HCV) is the major etiologic agent of transfusion non-A, non-B hepatitis. The NS3 protein has the typical amino acid motifs of the DEAD box family, the RNA helicase superfamily among the nonstructural proteins of HCV. The characterization of the detailed biochmical properties of the HCV NS3 protein, which seems to be a component of the replication complex, was necessary for understanding of the mechanism of HCV replication. The RNA helicase activity of the HCV NS3 protein was previously determined. To compare the biochemical properties of the HCV NS3 protein with the other helicases, and to assign the in vivo function of this helicase in the cellular and viral life cycle from the comparative data, the detailed characterization of the RNA binding and helicase activities was performed. The preference for the poly(U) and poly(A) in the RNA binding reaction implies that the HCV NS3 protein could easily bind to the 3````tail of the HCV RNA genome and begin the replication of the viral genome from the tail. The HCV RNA helicase unwinds RNA/DNA heteroduplexes as well as RNA/RNA duplexes in the aspects of the substrate specificity, and it catalytically translocates in the 3```` to 5```` direction. The HCV NS3 protein could unwind the DNA duplex as well. This DNA helicase activity implies that the HCV NS3 protein may be involved in the transcription, repair, and replication of the infected cell, as well as the viral gemone replication, like the other multifunctional RNA and DNA helicases. To assign the mechanism of this helicase, the detailed biochemical analysis on the DNA helicase activity was performed. Th e high affinity for the ATP analogs and the low affinity for ADP imply the mechanism of the helicase reaction; when ATP bound to the HCV NS3 protein is hydrolyzed, ADP can be quickly dissociated. The results in the competitive experiments suggest that the NS3 protein does not have dsRNA and ssDNA binding region. The high processivity (up to 50...