Cooperative translocation enhances the unwinding of duplex DNA by SARS coronavirus helicase nsP13

Lee, Na-Ra; Kwon, Hyun-Mi; Park, Kkothanahreum; Oh, Sangtaek; Jeong, Yong-Joo; Kim, Dong-Eun

Based on the results obtained when the length of the 5 0 -ssDNA tail and internal ssDNA gap in duplex DNA was varied, together with the observation that nsP13 oligomerization occurs in the presence of ssDNA, we believe that nsP13 more efficiently unwinds partial-duplex nucleic acid substrates containing ss regions at the 5 0 -side that are long enough to allow the binding of multiple monomers (Figure 7 ). The proposed model, schematically shown in Figure 7 , is very similar to that proposed for the unwinding of DNA by the HCV NS3 helicase and T4 Dda helicase (26, 41) . The proposed model for DNA unwinding by those helicases does not require protein oligomerization per se to obtain greater processivity. The increase in the helicase activity was explained by functional cooperativity rather than structural contacts between helicase monomers. Although multiple nsP13 helicases do not necessarily need to form a functional oligomeric form, multiple molecules of nsP13 are more efficient at unwinding longer substrates under single-turnover conditions. As depicted in Figure 7 , multiple molecules of nsP13 loaded on the leading strand are more likely to complete the unwinding process before leading helicase monomers disassociate from the substrate. This cooperative translocation is demonstrated by our results on the unwinding of the gapped duplex DNAs with or without the 5 0 -ssDNA overhang ( Figure 5 ). Unwinding of the 3 0 -side of the DNA duplex with a short gap size was impeded when fewer nsP13 molecules translocated from the 5 0 -side of the DNA, suggesting that multiple nsP13 monomers loaded on the same ssDNA track function together to produce more displaced ssDNA than a single nsP13 monomer can do. Taken together, the model presented in Figure 7 is a summary of how the non-processive nsP13 belonging to SF 1 helicase accomplishes substantial processivity in unwinding duplex nucleic acids in vitro. It should be noted that nsP13 may interact with other hosts and/or viral proteins when unwinding its RNA genome in infected cells and it may exhibit different properties during RNA unwinding.