The Hepatitis C Virus (HCV) NS3 protein contains amino acid motifs of a serine proteinase, a nucleotide triphosphatase (NTPase), and an RNA helicase. A carboxy fragment of the HCV NS3 protein was purified and possessed RNA helicase activity. Detections from the amino terminus resulted in the protein becoming soluble. Deletions from the carboxy terminus do not result in a loss of helicase activity until at least 50 amino acids are deleted. The helicase activity requires ATP and divalent cations such as Mg²⁺ and Mn²⁺. The helicase activity was blocked by monoclonal antibody specific to the HCV NS3 protein.