Role of the Conserved DECH-box Cysteine in Coupling Hepatitis C Virus Helicase-catalyzed ATP hydrolysis to RNA Unwinding

DECH-box proteins are a subset of DExH/D-box superfamily 2 helicases possessing a conserved Asp-Glu-Cys-His motif in their ATP binding site. The conserved His helps position the Asp and Glu residues, which coordinate the divalent metal cation that connects the protein to ATP and activate the water molecule needed for ATP hydrolysis, but the role of the Cys is still unclear. This study uses site-directed mutants of the model DECH-box helicase encoded by the hepatitis C virus (HCV) to examine the role of the Cys in helicase action. Proteins lacking a Cys unwound DNA less efficiently than wild-type proteins did. For example, at low protein concentrations, a helicase harboring a Gly instead of the DECH-box Cys unwound DNA more slowly than the wild-type helicase did, but at higher protein concentrations, the two proteins unwound DNA at similar rates. All HCV proteins analyzed had similar affinities for ATP and nucleic acids and hydrolyzed ATP in the presence of RNA at similar rates. However, in the absence of ...