Structural and functional studies of novel mechanisms of Lassa fever virus nucleoprotein in immune suppression, viral RNA transcription and replication
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Lassa fever virus is one of the most dangerous viruses of arenaviridae family, causing more than 500,000 infections per year in Africa. The fatality rate for hospitalized patients is as high as 20%. Due to the high fatality and lack of efficient licensed drugs and vaccines to treat and prevent, Lassa fever virus is classified as a Category A priority pathogen and biosafety level-4 agent by the Centers for Disease Control and Prevention of the USA. Cases were also found in the Americas and European countries, highlighting its potency to be a bioterrorism weapon. Like other areanaviruses, Lassa virus has developed a unique interferon suppression mechanism to evade from the host immune system, in which Lassa nucleoprotein plays the key role. To understand the LASV nucleoprotein functions, we tried to determine the first arenaviral nucleoprotein structure, LASV nucleoprotein. The LASV nucleoprotein (NP) was overexpressed and purified. The NP protein was crystallized and the structure was determined to 1.80 Å resolution. The crystals belong to space group P3, with the unit cell parameters a = b = 177.16 Å, c = 56.49 Å, α= β= 90° and γ= 120°. The LASV NP structure contains two domains, which are not similar to any reported viral nucleoprotein structures. The N-terminal domain has a novel structure with a cavity, which we proposed for cap binding, and the C-terminus is a 3’-5' ribonuclease, which is responsible for suppressing interferon production. To characterize the possible interaction between NP and other arenaviral protein, we also overexpressed and purified LASV Z. Interestingly, both NP and Z proteins have two forms and the purified NP protein and monomeric Z protein bind RNA. It is surprising that only the oligomeric Z protein interacts with NP protein but the monomeric Z protein does not as determined by Isothermal Titration Calorimetry (ITC). Our studies have reported the first arenaviral nucleoprotein structure, revealed the novel mechanism for the cap binding and immune suppression, which set up a platform for the development of novel drugs and vaccines to treat deadly arenaviral infections.
Thesis, PhD Doctor of Philosophy