Nanobody-mediated neutralization reveals an achilles heel for norovirus

• Verfasst von: Koromyslova, Anna D. [VerfasserIn]
• Verfasst von: Devant, Jessica [VerfasserIn]
• Verfasst von: Kiliç, Turgay [VerfasserIn]
• Verfasst von: Sabin, Charles [VerfasserIn]
• Verfasst von: Malak, Virginie [VerfasserIn]
• Verfasst von: Hansman, Grant S. [VerfasserIn]
• Titel: Nanobody-mediated neutralization reveals an achilles heel for norovirus
• Verf.angabe: Anna D. Koromyslova, Jessica M. Devant, Turgay Kilic, Charles D. Sabin, Virginie Malak, Grant S. Hansman
• E-Jahr: 2020
• Jahr: 16 June 2020
• Fussnoten: Gesehen am 27.08.2020
• Titel Quelle: Enthalten in: Journal of virology
• Ort Quelle: Baltimore, Md. : Soc., 1967
• Jahr Quelle: 2020
• Band/Heft Quelle: 94(2020,13) Artikel-Nummer 0660-20, 22 Seiten
• ISSN Quelle: 1098-5514
• DOI: doi:10.1128/JVI.00660-20
• Datenträger: Online-Ressource
• Sprache: eng
• K10plus-PPN: 1727891309

Abstract

Human norovirus frequently causes outbreaks of acute gastroenteritis. Although discovered more than five decades ago, antiviral development has, until recently, been hampered by the lack of a reliable human norovirus cell culture system. Nevertheless, a lot of pathogenesis studies were accomplished using murine norovirus (MNV), which can be grown routinely in cell culture. In this study, we analyzed a sizeable library of nanobodies that were raised against the murine norovirus virion with the main purpose of developing nanobody-based inhibitors. We discovered two types of neutralizing nanobodies and analyzed the inhibition mechanisms using X-ray crystallography, cryo-electron microscopy (cryo-EM), and cell culture techniques. The first type bound on the top region of the protruding (P) domain. Interestingly, this nanobody binding region closely overlapped the MNV receptor-binding site and collectively shared numerous P domain-binding residues. In addition, we showed that these nanobodies competed with the soluble receptor, and this action blocked virion attachment to cultured cells. The second type bound at a dimeric interface on the lower side of the P dimer. We discovered that these nanobodies disrupted a structural change in the capsid associated with binding cofactors (i.e., metal cations/bile acid). Indeed, we found that capsids underwent major conformational changes following addition of Mg2+ or Ca2+. Ultimately, these nanobodies directly obstructed a structural modification reserved for a postreceptor attachment stage. Altogether, our new data show that nanobody-based inhibition could occur by blocking functional and structural capsid properties. - IMPORTANCE This research discovered and analyzed two different types of MNV-neutralizing nanobodies. The top-binding nanobodies sterically inhibited the receptor-binding site, whereas the dimeric-binding nanobodies interfered with a structural modification associated with cofactor binding. Moreover, we found that the capsid contained a number of vulnerable regions that were essential for viral replication. In fact, the capsid appeared to be organized in a state of flux, which could be important for cofactor/receptor-binding functions. Blocking these capsid-binding events with nanobodies directly inhibited essential capsid functions. Moreover, a number of MNV-specific nanobody binding epitopes were comparable to human norovirus-specific nanobody inhibitors. Therefore, this additional structural and inhibition information could be further exploited in the development of human norovirus antivirals.