Figure 6.

Structural Basis for Possible nsp13 Helicase Functions during Viral Genome Replication-Transcription

Structural models are shown as cartoons (holo-RdRp, light blue; nsp13.1 helicase, orange shades; RNA strands, colored tubes). The nsp13.2 helicase is not shown for clarity (all models are compatible with the presence of nsp13.2). With each structural diagram is a schematic cartoon illustrating the arrangement of RNA strands. Additional proteins involved in these processes are omitted. The product RNA (p-RNA) being elongated by the RdRp is shown in red.

(A) The SARS-CoV-2 nsp13-RdRp cryo-EM structure likely represents an equilibrium between two states.

(B) During RNA synthesis on a single-stranded RNA template (cyan), nsp13 could function distributively to clear downstream RNA secondary structure (or RNA binding proteins). Similarly, on a duplex RNA template (cyan and green), nsp13 could processively unwind downstream duplex RNA.

(C) Proposed helicase function during template switching associated with sub-genomic transcription (sg-transcription) (Enjuanes et al., 2006; Lehmann et al., 2015b; Pasternak et al., 2001; Snijder et al., 2016; Sola et al., 2015). (i) Negative-strand RNA synthesis proceeds from the genomic 3′ poly(A)-tail until a transcription-regulating sequence (TRS-R, orange) (Alonso et al., 2002) is transcribed (cTRS, yellow). (ii) The TRS causes transcription complex stalling. (iii) Helicase function acting on the + strand RNA (cyan) causes backtracking of the transcription complex, freeing the pRNA 3′ end. (iv) The p-RNA 3′-end cTRS (yellow) hybridizes with the complementary TRS-L (orange) following the genomic 5′ leader sequence (magenta) (Alonso et al., 2002; Pasternak et al., 2001; Zúñiga et al., 2004). (v) Processive helicase function backtracks the RdRp complex and unwinds the p-RNA from the genomic 3′ end. A second RdRp complex (holo-RdRp2) can load into the p-RNA 3′ end and continue transcription using the 5′ leader as a template.