Myxovirus resistance (Mx) GTPases are induced by interferons and inhibit multiple viruses including influenza, Thogoto virus (THOV), and the human immunodeficiency viruses. They have the characteristic domain architecture of dynamin-related proteins with an amino-terminal GTPase (G) domain, a bundle signaling element (BSE), and a carboxy-terminal stalk responsible for self-assembly and effector functions. Human MxA is localized in the cytoplasm and is partly associated with membranes of the smooth endoplasmic reticulum (ER). It shows a protein concentration-dependent increase in GTPase activity, indicating regulation of GTP hydrolysis via G domain dimerization. However, the exact mechanism of GTP hydrolysis and the function of GTP binding and hydrolysis for the antiviral activity have not been characterized so far. To clarify the role of GTP binding and the importance of the G domain interface for the catalytic and antiviral function of MxA, I performed a thorough biochemical characterization in the first part of this doctoral thesis. Based on structure-based mutagenesis, residues crucial for nucleotide-binding and dimerization were analyzed. The closely related human MxB protein is a potent restriction factor for HIV-1 and other lentiviruses, in addition to its already known involvement in regulating nucleocytoplasmic transport and cell-cycle progression. However, the role of GTP binding and hydrolysis in restricting HIV-1 is still under debate. Consequently, I analyzed the GTPase activity of MxB in absence and presence of MxA to deduce possible differences to MxA. A biochemical characterization of MxB might shed more light on their differential antiviral spectrum. The tick-borne transmitted THOV NP was identified as a target of the MxA GTPase. In viral particles, the NP is encapsulating the viral RNA, and together with the viral polymerase, they form the viral ribonucleoparticles (vRNPs), which are essential for the transcription and replication of the virus. The second part of this thesis focuses on a basic biochemical characterization of the THOV and DHOV NP to understand the mechanism of RNA binding and oligomerization. To better understand the structural features of orthomyxovirus RNP formation, I solved the atomic structure of the THOV NP. Finally, the atomic model of the THOV NP helps to identify putative interaction sites with the human MxA protein, and provides a structural model for other orthomyxoviruses, including influenza viruses.
