Characterization of the hepatitis C viral helicase, and, a sequence analysis algorithm that predicts functional interactions

The Hepatitis C Virus (HCV) is a cytoplasmically replicating RNA virus that is a major worldwide health problem. The Hepatitis C viral protein NS3 is a protease/helicase that plays an essential role in the life cycle of this virus. NS3 is therefore an important drug target in the effort to combat HCV. Most work, however, has focused on the protease, rather than the helicase activities of the enzyme. In order to better characterize NS3 helicase activity, we evaluated individual stages of duplex unwinding by NS3 alone and in complex with cofactor NS4A. Despite a putative replicative role in RNA unwinding, we found that NS3 alone is a surprisingly poor helicase on RNA, but that RNA activity is promoted by cofactor NS4A. In contrast, NS3 alone is a highly processive helicase on DNA. Phylogenctic analysis suggests that this robust DNA helicase activity is not vestigial, and may have specifically evolved in HCV. Given that HCV has no replicative DNA intermediate, these findings suggest that NS3 may have the capacity to affect host DNA.; Additionally, a novel, automated sequence analysis algorithm has been designed that is capable of predicting function interactions between residues. This algorithm is uniquely applicable to any type of biopolymer, including RNAs and proteins, as well as combinations of different biopolymers. This algorithm, entitled SHEVEK, yields accurate results despite using alignments of only a small number of sequences. The versatility and accuracy of the algorithm is demonstrated by the prediction of interactions within RNAs and modified RNAs, within proteins, as well as through the prediction of RNA-protein and protein-protein interactions.