Much of our work has focused on two viral DExH proteins that are essential for the replication of pox viruses and hepatitis C (HCV) viruses. These helicases serve as model systems for defining the behavior of DExH/D proteins in general, and more specifically, they are important drug targets in the effort to develop antiviral therapeutics. Using transient kinetics and chemogenetic approaches, we have examined the mechanism by which these proteins translocate, unwind nucleic acid, and strip proteins from RNA strands.

In studies of the NPH-II protein from vaccinia virus we linked the extent of RNA unwinding (e.g. work) with the utilization of ATP, thereby demonstrating that DExH/D proteins are true molecular motors that act on RNA. NPH-II is a processive, directional motor that unwinds RNA with a kinetic step-size of 6 base-pairs and a quantifiable translocation rate constant that depends on the nature of the metal ion cofactor (Jankowsky & Pyle et al, Nature 2000). By examining NPH-II unwinding of chemically modified RNA substrates, we have shown that the protein tracks along the sugar-phosphate backbone of the substrate loading strand while stripping away the top strand without regard to its chemical identity (see papers by Kawaoka & Pyle et al). Other types of objects can also be displaced from the loading strand, as we demonstrated that NPH-II can actively displace proteins from RNA in an ATP-dependent manner (Jankowsky & Pyle, Science 2001). This finding established the precedent for a new enzymatic activity (termed RNPase function), that is now widely hypothesized to be important for the function of macromolecular machines such as the spliceosome (Tanner & Linder, Mol Cell 2001; Fairman & Jankowsky, Science 2004). In these and other experiments, the NPH-II helicase continues to serve as an indispensable model system for exploring the scope of mechanical capabilities inherent to SF2 proteins.