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

A plot of the final reaction amplitude versus duplex length showed that processivity of the nsP13 helicase in unwinding of DNA with a 5 0 -tail of 20 nt is very low ( Figure 2B ) compared with other related helicases, such as NPH-II RNA helicase and HCV NS3 DNA helicase (33, 34) . The processivity of a helicase is defined as the probability of unwinding a base pair versus dissociation of the helicase from the DNA at that position (24) . At present, it is hard to calculate the exact processivity of the nsP13 helicase due to the absence of 'kinetic step size' information of active nsP13 (31, 35) . The monomeric form of most SF1 and SF2 helicases shows that they are rapid and processive at translocating along ssDNA, but not processive in terms of dsDNA unwinding (36) (37) (38) . This raises the possibility that nsP13 may function as an oligomer in its active form like other SF1 and SF2 helicases, despite the fact that the helicase is nonprocessive by itself. Considering the hexameric ringshaped helicases such as T7 bacteriophage helicase, the formation of ring was expected to result in high processivity due to DNA passage through a central channel. However, single-turnover kinetic analysis showed that the T7 helicase by itself has a low processivity for unwinding dsDNA ($15 bp), but had a 9-fold higher helicase activity ($130 bp) in the presence of polymerase and thioredoxin (31, 39) . The fact that SARS coronavirus has a polymerase and many non-structural proteins of unknown functions led to the hypothesis that additional proteins may be involved in the helicase activity of nsP13 (40) .