title: Biochemical and biophysical characterization of cell-free synthesized Rift Valley fever virus nucleoprotein capsids enables in vitro screening to identify novel antivirals.
abstract: Viral capsid assembly involves the oligomerization of the capsid nucleoprotein (NP), which is an essential step in viral replication and may represent a potential antiviral target. An in vitro transcription-translation reaction using a wheat germ (WG) extract in combination with a sandwich ELISA assay has recently been used to identify small molecules with antiviral activity against the rabies virus.
  
  
  
  Here, we examined the application of this system to viruses with capsids with a different structure, such as the Rift Valley fever virus (RVFV), the etiological agent of a severe emerging infectious disease. The biochemical and immunological characterization of the in vitro-generated RVFV NP assembly products enabled the distinction between intermediately and highly ordered capsid structures. This distinction was used to establish a screening method for the identification of potential antiviral drugs for RVFV countermeasures.
  
  
  
  These results indicated that this unique analytical system, which combines nucleoprotein oligomerization with the specific immune recognition of a highly ordered capsid structure, can be extended to various viral families and used both to study the early stages of NP assembly and to assist in the identification of potential antiviral drugs in a cost-efficient manner.
  
  
  
  Reviewed by Jeffry Skolnick and Noah Isakov. For the full reviews please go to the Reviewers' comments section.
authors.affiliation: Prosetta Antiviral Inc, San Francisco, CA, USA.
authors.forename: Sean
authors.initials: S
authors.lastname: Broce
authors.affiliation: NIAID, Ft Detrick, MD, USA.
authors.forename: Lisa
authors.initials: L
authors.lastname: Hensley
authors.affiliation: Prosetta Antiviral Inc, San Francisco, CA, USA.|DuPont Industrial Biosciences, Palo Alto, CA, USA.
authors.forename: Tomoharu
authors.initials: T
authors.lastname: Sato
authors.affiliation: Prosetta Antiviral Inc, San Francisco, CA, USA.|Global Blood Therapeutics, Inc, Palo Alto, CA, USA.
authors.forename: Joshua
authors.initials: J
authors.lastname: Lehrer-Graiwer
authors.affiliation: Prosetta Antiviral Inc, San Francisco, CA, USA.
authors.forename: Christian
authors.initials: C
authors.lastname: Essrich
authors.affiliation: CUBRC, Inc, Buffalo, NY, USA.
authors.forename: Katie J
authors.initials: KJ
authors.lastname: Edwards
authors.affiliation: Life Technologies, Inc, Grand Island, NY, USA.
authors.forename: Jacqueline
authors.initials: J
authors.lastname: Pajda
authors.affiliation: Biomedical Advanced Research & Development Authority, Office of the Assistant Secretary of Preparedness & Response, U.S. Department of Health & Human Services, Washington, DC, USA.
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Davis
authors.affiliation: Prosetta Antiviral Inc, San Francisco, CA, USA.|Biocon Bristol-Myers Squibb, Syngene International Ltd, Bangalore, India.
authors.forename: Rami
authors.initials: R
authors.lastname: Bhadresh
authors.affiliation: Prosetta Antiviral Inc, San Francisco, CA, USA.
authors.forename: Clarence R
authors.initials: CR
authors.lastname: Hurt
authors.affiliation: Prosetta Antiviral Inc, San Francisco, CA, USA.
authors.forename: Beverly
authors.initials: B
authors.lastname: Freeman
authors.affiliation: Prosetta Antiviral Inc, San Francisco, CA, USA.
authors.forename: Vishwanath R
authors.initials: VR
authors.lastname: Lingappa
authors.affiliation: Prosetta Antiviral Inc, San Francisco, CA, USA.|Bigelow Aerospace Advanced Space Studies, Las Vegas, NV, USA.
authors.forename: Colm A
authors.initials: CA
authors.lastname: Kelleher
authors.affiliation: Prosetta Antiviral Inc, San Francisco, CA, USA. mvkarpuj@bioa2z.com.|CUBRC, Inc, Buffalo, NY, USA. mvkarpuj@bioa2z.com.|BioA2Z, Inc, San Francisco, CA, USA. mvkarpuj@bioa2z.com.|Faculty of Medicine, Bar-Ilan University, 8 Henrietta Szold, Safed, 1311502, Israel. mvkarpuj@bioa2z.com.
authors.forename: Marcela V
authors.initials: MV
authors.lastname: Karpuj
mesh_terms: D000998:Antiviral Agents / Q000032:analysis*; D002213:Capsid / Q000502:physiology*; D002474:Cell-Free System; D055808:Drug Discovery / Q000379:methods*; D004353:Drug Evaluation, Preclinical; D009698:Nucleoproteins / Q000737:chemistry; D012296:Rift Valley fever virus / Q000502:physiology*
chemical_list: D000998:Antiviral Agents; D009698:Nucleoproteins
keywords: Antivirals; Capsid nucleoprotein oligomerization; Cell-free protein synthesis; Rift Valley fever virus; Wheat germ extracts