Leung Daisy W, Ginder Nathaniel D, Fulton D Bruce, Nix Jay, Basler Christopher F, Honzatko Richard B, Amarasinghe Gaya K
Department of Biochemistry, Biophysics and Molecular Biology, Iowa State University, Ames, IA 50011, USA.
Proc Natl Acad Sci U S A. 2009 Jan 13;106(2):411-6. doi: 10.1073/pnas.0807854106. Epub 2009 Jan 2.
Ebola viruses (EBOVs) cause rare but highly fatal outbreaks of viral hemorrhagic fever in humans, and approved treatments for these infections are currently lacking. The Ebola VP35 protein is multifunctional, acting as a component of the viral RNA polymerase complex, a viral assembly factor, and an inhibitor of host interferon (IFN) production. Mutation of select basic residues within the C-terminal half of VP35 abrogates its dsRNA-binding activity, impairs VP35-mediated IFN antagonism, and attenuates EBOV growth in vitro and in vivo. Because VP35 contributes to viral escape from host innate immunity and is required for EBOV virulence, understanding the structural basis for VP35 dsRNA binding, which correlates with suppression of IFN activity, is of high importance. Here, we report the structure of the C-terminal VP35 IFN inhibitory domain (IID) solved to a resolution of 1.4 A and show that VP35 IID forms a unique fold. In the structure, we identify 2 basic residue clusters, one of which is important for dsRNA binding. The dsRNA binding cluster is centered on Arg-312, a highly conserved residue required for IFN inhibition. Mutation of residues within this cluster significantly changes the surface electrostatic potential and diminishes dsRNA binding activity. The high-resolution structure and the identification of the conserved dsRNA binding residue cluster provide opportunities for antiviral therapeutic design. Our results suggest a structure-based model for dsRNA-mediated innate immune antagonism by Ebola VP35 and other similarly constructed viral antagonists.
埃博拉病毒（EBOV）可引发人类罕见但致死率极高的病毒性出血热疫情，目前针对这些感染尚无获批的治疗方法。埃博拉病毒VP35蛋白具有多种功能，它是病毒RNA聚合酶复合体的组成部分、病毒组装因子以及宿主干扰素（IFN）产生的抑制剂。VP35 C端结构域内特定碱性残基的突变会消除其双链RNA（dsRNA）结合活性，损害VP35介导的IFN拮抗作用，并在体外和体内减弱埃博拉病毒的生长。由于VP35有助于病毒逃避宿主固有免疫，且是埃博拉病毒毒力所必需的，因此了解与IFN活性抑制相关的VP35 dsRNA结合的结构基础至关重要。在此，我们报告了C端VP35 IFN抑制结构域（IID）的结构，其分辨率为1.4 Å，并表明VP35 IID形成了独特的折叠结构。在该结构中，我们鉴定出2个碱性残基簇，其中一个对dsRNA结合很重要。dsRNA结合簇以Arg-312为中心，这是IFN抑制所需的高度保守残基。该簇内残基的突变会显著改变表面静电势并降低dsRNA结合活性。高分辨率结构以及保守dsRNA结合残基簇的鉴定为抗病毒治疗设计提供了机会。我们的结果提出了一种基于结构的模型，用于解释埃博拉病毒VP35和其他结构类似的病毒拮抗剂介导的dsRNA先天免疫拮抗作用。
