Characterization of the Zika virus two-component NS2B-NS3 protease and structure-assisted identification of allosteric small-molecule antagonists

Antiviral Res. 2017 Jul:143:218-229. doi: 10.1016/j.antiviral.2017.04.015. Epub 2017 Apr 29.


The recent re-emergence of Zika virus (ZIKV)1, a member of the Flaviviridae family, has become a global emergency. Currently, there are no effective methods of preventing or treating ZIKV infection, which causes severe neuroimmunopathology and is particularly harmful to the developing fetuses of infected pregnant women. However, the pathology induced by ZIKV is unique among flaviviruses, and knowledge of the biology of other family members cannot easily be extrapolated to ZIKV. Thus, structure-function studies of ZIKV proteins are urgently needed to facilitate the development of effective preventative and therapeutic agents. Like other flaviviruses, ZIKV expresses an NS2B-NS3 protease, which consists of the NS2B cofactor and the NS3 protease domain and is essential for cleavage of the ZIKV polyprotein precursor and generation of fully functional viral proteins. Here, we report the enzymatic characterization of ZIKV protease, and we identify structural scaffolds for allosteric small-molecule inhibitors of this protease. Molecular modeling of the protease-inhibitor complexes suggests that these compounds bind to the druggable cavity in the NS2B-NS3 protease interface and affect productive interactions of the protease domain with its cofactor. The most potent compound demonstrated efficient inhibition of ZIKV propagation in vitro in human fetal neural progenitor cells and in vivo in SJL mice. The inhibitory scaffolds could be further developed into valuable research reagents and, ultimately, provide a roadmap for the selection of efficient inhibitors of ZIKV infection.

Keywords: Flavivirus; Inhibitors; NS2B; NS3; Protease; Zika virus.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Allosteric Site*
  • Amino Acid Sequence
  • Animals
  • Antiviral Agents / antagonists & inhibitors
  • Antiviral Agents / chemistry
  • Base Sequence
  • Enzyme Activation
  • Female
  • Flavivirus / chemistry
  • Gene Expression
  • Humans
  • Inhibitory Concentration 50
  • Mice
  • Models, Molecular
  • Protease Inhibitors / chemistry*
  • Protease Inhibitors / pharmacology*
  • Protein Binding
  • Protein Conformation
  • Protein Interaction Domains and Motifs
  • RNA Helicases / chemistry
  • RNA Helicases / drug effects
  • SOXB1 Transcription Factors / genetics
  • Sequence Alignment
  • Serine Endopeptidases / chemistry
  • Serine Endopeptidases / drug effects
  • Stem Cells
  • Viral Nonstructural Proteins / chemistry*
  • Viral Nonstructural Proteins / drug effects
  • Viral Proteins / chemistry
  • Viral Proteins / genetics
  • Zika Virus / chemistry
  • Zika Virus / enzymology*
  • Zika Virus / genetics
  • Zika Virus / growth & development
  • Zika Virus Infection / virology


  • Antiviral Agents
  • NS2B protein, flavivirus
  • NS3 protein, flavivirus
  • Protease Inhibitors
  • SOX2 protein, human
  • SOXB1 Transcription Factors
  • Viral Nonstructural Proteins
  • Viral Proteins
  • Serine Endopeptidases
  • RNA Helicases