The SARS-coronavirus nsp7+nsp8 complex is a unique multimeric RNA polymerase capable of both de novo initiation and primer extension

te Velthuis, Aartjan J.W.; van den Worm, Sjoerd H. E.; Snijder, Eric J.

Comparing our data against the background of the previously published nsp(7+8) structure (13), we made four main observations. Firstly, we note that in the published nsp(7+8) crystal structure four of the eight N-terminal D/ExD/E motifs in the complex reside at the border of partially unresolved N-terminal nsp8 domains, where the coordinates of up to 49 nsp8 residues and 5 exogenous amino acids derived from the removed GST fusion partner were not determined. In light of our own finding that unnatural N-terminal extensions severely impair nsp8's RdRp activity ( Figure 5 ), this suggests that the published crystal structure may not represent an active conformation of the nsp(7+8) polymerase. Secondly, we observe that residues D50 and D52, which are both crucial for nsp(7+8) activity, are residing in an a-helix in the nsp(7+8) structure (Supplementary Figure S5) , whereas in canonical primases and polymerases, the catalytic centre is preferentially located on b-strands or turns (12, 22) . Thirdly, we note that Mg 2+ was lacking from the published nsp(7+8) crystal structure (13) , even though it is required for nsp(7+8) activity. Fourthly and last, we observe that a 1:1 ratio of nsp7:nsp8 is sufficient to capture all nsp8 in a higher molecular weight complex ( Figure 2F ) whereas previously a 2:1 ratio was required (13) , potentially due to the additional N-terminal residues that altered the dynamics of complex formation.