Characterization and applications of a Crimean-Congo hemorrhagic fever virus nucleoprotein-specific Affimer: Inhibitory effects in viral replication and development of colorimetric diagnostic tests.
[en] Crimean-Congo hemorrhagic fever orthonairovirus (CCHFV) is one of the most widespread medically important arboviruses, causing human infections that result in mortality rates of up to 60%. We describe the selection of a high-affinity small protein (Affimer-NP) that binds specifically to the nucleoprotein (NP) of CCHFV. We demonstrate the interference of Affimer-NP in the RNA-binding function of CCHFV NP using fluorescence anisotropy, and its inhibitory effects on CCHFV gene expression in mammalian cells using a mini-genome system. Solution of the crystallographic structure of the complex formed by these two molecules at 2.84 Å resolution revealed the structural basis for this interference, with the Affimer-NP binding site positioned at the critical NP oligomerization interface. Finally, we validate the in vitro application of Affimer-NP for the development of enzyme-linked immunosorbent and lateral flow assays, presenting the first published point-of-care format test able to detect recombinant CCHFV NP in spiked human and animal sera.
Disciplines :
Microbiology
Author, co-author :
Álvarez-Rodríguez, Beatriz ; School of Molecular and Cellular Biology, University of Leeds, Leeds, United Kingdom
Tiede, Christian ; School of Molecular and Cellular Biology, University of Leeds, Leeds, United Kingdom
Hoste, Alexis ; Université de Liège - ULiège > Département GxABT > Microbial technologies ; School of Molecular and Cellular Biology, University of Leeds, Leeds, United Kingdom ; INGENASA, Inmunología y Genética Aplicada S.A., Madrid, Spain
Surtees, Rebecca A; School of Molecular and Cellular Biology, University of Leeds, Leeds, United Kingdom
Trinh, Chi H; School of Molecular and Cellular Biology, University of Leeds, Leeds, United Kingdom
Slack, Gillian S; National Infection Service, Public Health England, Porton Down, Salisbury, United Kingdom
Chamberlain, John ; National Infection Service, Public Health England, Porton Down, Salisbury, United Kingdom
Hewson, Roger; National Infection Service, Public Health England, Porton Down, Salisbury, United Kingdom
Fresco, Alba ; INGENASA, Inmunología y Genética Aplicada S.A., Madrid, Spain
Sastre, Patricia; INGENASA, Inmunología y Genética Aplicada S.A., Madrid, Spain
Tomlinson, Darren C ; School of Molecular and Cellular Biology, University of Leeds, Leeds, United Kingdom
Millner, Paul A; School of Molecular and Cellular Biology, University of Leeds, Leeds, United Kingdom
Edwards, Thomas A; School of Molecular and Cellular Biology, University of Leeds, Leeds, United Kingdom
Barr, John N ; School of Molecular and Cellular Biology, University of Leeds, Leeds, United Kingdom
Characterization and applications of a Crimean-Congo hemorrhagic fever virus nucleoprotein-specific Affimer: Inhibitory effects in viral replication and development of colorimetric diagnostic tests.
H2020 - 721367 - HONOURs - Host switching pathogens, infectious outbreaks and zoonosis; a Marie Sklodowska-Curie Training Network
Funders :
EU - European Union
Funding text :
This work was supported by an EU Marie Skłodowska-Curie Actions (MSCA, ec.europa.eu) Innovative Training Network (ITN): H2020-MSCA-ITN-2016, under grant No 721367 (to B. Álvarez-Rodríguez). The funders had no role in study.
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