Phage display of the Equine arteritis virus nsp1 ZF domain and examination of its metal interactions

Martin B. Oleksiewicz, E.J. Snijder, Preben Normann

    Research output: Contribution to journalJournal articleResearchpeer-review


    A putative zinc finger (ZF) domain in the Equine arteritis virus (EAV) nsp 1 protein was described recently to be required for viral transcription. The nsp 1 ZF (50 aa) was expressed on the surface of M13KE gIII phage, fused to the N terminus of the phage pIII protein. To evaluate the functionality of the ZF domain, a binding assay was developed, based on the use of immobilized Ni2+ ions (Ni-NTA). Phages displaying ZF bound significantly better to Ni-NTA than did phages displaying negative-control peptides, which also contained metal-coordinating residues. Also, binding of ZF-displaying phages could be inhibited by an anti-nsp 1 serum, or by mutation of residues predicted to be important for zinc coordination. Finally, binding was abolished by low concentrations (0.1%) Tween 20, and rescued by including Zn2+, Ni2+ or Cu2+, but not Mg2+, in the binding buffer, suggesting that formation of secondary structure was involved in binding of the ZF to Ni-NTA. These findings provide the first experimental evidence that the putative nsp 1 ZF domain can coordinate divalent metal ions, and that this property is associated with the secondary structure of the domain. The Ni-NTA binding assay developed in the present study may have general applications in the study of other ZF domains.
    Original languageChinese
    JournalJournal of Virological Methods
    Issue number2
    Pages (from-to)159-169
    Publication statusPublished - 2004


    • IMAC
    • arterivirus
    • Nsp1
    • nickel
    • zinc finger
    • nidovirus
    • replicase

    Cite this