Characterization of nsP-specific nanobodies targeting Chikungunya and Semliki Forest Virus
Abstract: Viral infections are constantly increasing and impose a large threat to the public health. Alphaviruses are responsible for several animal and human diseases and have a large medical importance with few treatments available today. Alphaviruses are small, spherical single stranded RNA viruses, and are most often transmitted by mosquito vectors. Alphaviruses contains a domain of nonstructural proteins that compose the replication machinery. The domain is crucial for viral replication to occur and is therefore an interesting target for antiviral therapy. With the focus on Chikungunya and Semliki Forest Virus this work investigates the events in the cells on molecular level during infections. To do this a panel of Camelid derived single domain antibodies are developed to target the nonstructural proteins of Chikungunya and Semliki Forest Virus. Binding of the produced nanobodies to the viral proteins was investigated by biochemical methods including immunoprecipitations, western blot, and ELISA. Cell lines that express nsP-specific nanobodies in the cytosol were employed for infection- and plaque assays with Semliki Forest Virus in order to determine the antiviral potential of the new nanobodies. Three of the nanobodies proved to bind two different nonstructural proteins of the viruses, providing opportunities for further investigations and a possible use of these nanobodies to identify viral vulnerabilities that could be exploited for antiviral intervention.
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