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首页> 外文期刊>Nucleic Acids Research >Cooperative translocation enhances the unwinding of duplex DNA by SARS coronavirus helicase nsP13
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Cooperative translocation enhances the unwinding of duplex DNA by SARS coronavirus helicase nsP13

机译：合作易位通过SARS冠状病毒解旋酶nsP13增强双链DNA的解链

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摘要

SARS coronavirus encodes non-structural protein 13 (nsP13), a nucleic acid helicase/NTPase belonging to superfamily 1 helicase, which efficiently unwinds both partial-duplex RNA and DNA. In this study, unwinding of DNA substrates that had different duplex lengths and 5'-overhangs was examined under single-turnover reaction conditions in the presence of excess enzyme. The amount of DNA unwound decreased significantly as the length of the duplex increased, indicating a poor in vitro processivity. However, the quantity of duplex DNA unwound increased as the length of the single-stranded 5'-tail increased for the 50-bp duplex. This enhanced processivity was also observed for duplex DNA that had a longer single-stranded gap in between. These results demonstrate that nsP13 requires the presence of a long 5'-overhang to unwind longer DNA duplexes. In addition, enhanced DNA unwinding was observed for gapped DNA substrates that had a 5'-overhang, indicating that the translocated nsP13 molecules pile up and the preceding helicase facilitate DNA unwinding. Together with the propensity of oligomer formation of nsP13 molecules, we propose that the cooperative translocation by the functionally interacting oligomers of the helicase molecules loaded onto the 5'-overhang account for the observed enhanced processivity of DNA unwinding.
机译：SARS冠状病毒编码非结构蛋白13（nsP13），一种属于超家族1解旋酶的核酸解旋酶/ NTPase，可有效解旋部分双链RNA和DNA。在这项研究中，在存在过量酶的情况下，在单周转反应条件下检查了具有不同双链体长度和5'-突出端的DNA底物的解链。解链的DNA量随着双链体长度的增加而显着降低，表明体外的合成能力较差。但是，对于50 bp的双链体，解链的双链体DNA的数量随着单链5'尾巴长度的增加而增加。对于双链DNA，其间具有更长的单链间隙，也观察到了这种增强的合成能力。这些结果表明，nsP13需要长的5'突出端才能解开更长的DNA双链体。此外，观察到具有5'突出端的空缺DNA底物的DNA解链增强，表明易位的nsP13分子堆积并且前面的解旋酶促进DNA解链。连同nsP13分子形成寡聚体的倾向一起，我们提出通过加载到5'突出端的解旋酶分子的功能性相互作用寡聚体进行的合作易位，说明了观察到的DNA解链增强的生产力。

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