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

Enhancement of duplex DNA unwinding by increasing the length of 5 0 -overhang Partial duplex DNA substrates containing 50 bp with a 5 0 -ssDNA overhang of varying lengths (T n 50bp in Table 1 ) were designed in order to investigate whether DNA unwinding is dependent on the length of the loading strand. Although nsP13 has been shown to have stimulated ATPase activity in the presence of polynucleotides, detailed binding affinity experiments have not been conducted. More importantly, the extent of stimulation of ATPase activity with ss nucleic acids was previously shown to be proportional to the length of ssDNA or ssRNA at a given concentration (25) . These observations may indicate that nsP13 binds to the ss nucleic acids with a defined binding site size. Thus, we can speculate that as the length of the loading strand increases, more nsP13 helicases would be bound to the substrate DNA under the single-turnover reaction condition, in which the helicase outnumbers the amount of substrate DNAs.