Abstract
Non-structural proteins (nsp) constitute the SARS-CoV-2 replication and transcription complex (RTC) to play a pivotal role in the virus life cycle. Here we determine the atomic structure of a SARS-CoV-2 mini RTC, assembled by viral RNA-dependent RNA polymerase (RdRp, nsp12) with a template-primer RNA, nsp7 and nsp8, and two helicase molecules (nsp13-1 and nsp13-2), by cryo-electron microscopy. Two groups of mini RTCs with different conformations of nsp13-1 are identified. In both of them, nsp13-1 stabilizes overall architecture of the mini RTC by contacting with nsp13-2, which anchors the 5′-extension of RNA template, as well as interacting with nsp7-nsp8-nsp12-RNA. Orientation shifts of nsp13-1 results in its variable interactions with other components in two forms of mini RTC. The mutations on nsp13-1:nsp12 and nsp13-1:nsp13-2 interfaces prohibit the enhancement of helicase activity achieved by mini RTCs. These results provide an insight into how helicase couples with polymerase to facilitate its function in virus replication and transcription.
Highlights
Non-structural proteins constitute the SARS-CoV-2 replication and transcription complex (RTC) to play a pivotal role in the virus life cycle
The helicase activity of apo nsp[13] is enhanced by the formation of mini RTC (Fig. 1c), approving this mini RTC is functional, which is consistent with the previous result[8,11]
The results show that mini RTC with an nsp12R365A mutation has a compared helicase activity with the individual apo nsp[13], which is significantly lower than mini RTC
Summary
Non-structural proteins (nsp) constitute the SARS-CoV-2 replication and transcription complex (RTC) to play a pivotal role in the virus life cycle. We determine the atomic structure of a SARS-CoV-2 mini RTC, assembled by viral RNA-dependent RNA polymerase (RdRp, nsp12) with a template-primer RNA, nsp[7] and nsp[8], and two helicase molecules (nsp[] and nsp13-2), by cryo-electron microscopy. Two groups of mini RTCs with different conformations of nsp[] are identified In both of them, nsp[] stabilizes overall architecture of the mini RTC by contacting with nsp[], which anchors the 5′-extension of RNA template, as well as interacting with nsp7-nsp8-nsp12-RNA. SARS-CoV-2 encodes 16 non-structural proteins (nsp) to assemble a so-called replication and transcription complex (RTC) to facilitate virus replication and transcription[7]. We aim to dissect the mechanism for how helicase couples with central RTC
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