Structural basis for the regulatory function of a complex zinc-binding domain in a replicative arterivirus helicase resembling a nonsense-mediated mRNA decay helicase

Deng, Zengqin; Lehmann, Kathleen C.; Li, Xiaorong; Feng, Chong; Wang, Guoqiang; Zhang, Qi; Qi, Xiaoxuan; Yu, Lin; Zhang, Xingliang; Feng, Wenhai; Wu, Wei; Gong, Peng; Tao, Ye; Posthuma, Clara C.; Snijder, Eric J.; Gorbalenya, Alexander E.; Chen, Zhongzhou

The crystal structure of EAV nsp10" reveals a multi-domain organization of the arterivirus replicative helicase Because 3D structures of orthologous proteins were not available, we took advantage of the zinc-binding properties of nsp10 and used the zinc multiple-wavelength anomalous dispersion (MAD) method (42) to solve the EAV nsp10Á structure. The presence and position of three zinc atoms were established with anomalous data collected from the zinc absorption edge ( Table 1 ). The final model included EAV nsp10 residues 1-401, three zinc ions in the N-terminal ZBD, five sulphate ions and 267 water molecules. Two RecA-like a/b domains (1A and 2A) form the structure's C-terminal part ( Figure 2B ; cyan and green) and constitute the helicase core (HEL1). Domain 1A contains a parallel five-stranded b-sheet that is sandwiched by three a-helices on one side and two a-helices on the other. Domain 2A contains a parallel four-stranded b-sheet with five a-helices on the side facing domain 1A. Upstream of domain 1A, we identified an additional domain with a characteristic b-barrel fold (Figure 2A and B; magenta). It consists of five b-strands arranged as two tightly packed anti-parallel b-sheets and is juxtaposed to domain 1A ( Figure 2B ). The location of this domain in the protein sequence and its orientation relative to the HEL1 domain resemble those of domain 1B in helicases of the SF1B Upf1-like subfamily ( Figure 3B and C), and it was therefore named accordingly in our nsp10Á structure. The domain has no counterpart in the only other solved structure of a viral SF1 helicase, that from ToMV ( Figure 3A ) (15) , whereas its counterpart in helicases of the Pif1-like subfamily is inserted in domain 2A ( Figure 3D ) (48) .