Kolawole Abimbola O, Smith Hong Q, Svoboda Sophia A, Lewis Madeline S, Sherman Michael B, Lynch Gillian C, Pettitt B Montgomery, Smith Thomas J, Wobus Christiane E
Department of Microbiology and Immunology, University of Michigan Medical School, Ann Arbor, Michigan, USA.
Department of Biochemistry and Molecular Biology, University of Texas Medical Branch at Galveston, Galveston, Texas, USA.
mSphere. 2017 Oct 18;2(5). doi: 10.1128/mSphere.00334-17. eCollection 2017 Sep-Oct.
Ideal antiviral vaccines elicit antibodies (Abs) with broad strain recognition that bind to regions that are difficult to mutate for escape. Using 10 murine norovirus (MNV) strains and 5 human norovirus (HuNoV) virus-like particles (VLPs), we identified monoclonal antibody (MAb) 2D3, which broadly neutralized all MNV strains tested. Importantly, escape mutants corresponding to this antibody were very slow to develop and were distal to those raised against our previously studied antibody, A6.2. To understand the atomic details of 2D3 neutralization, we determined the cryo-electron microscopy (cryo-EM) structure of the 2D3/MNV1 complex. Interestingly, 2D3 binds to the top of the P domain, very close to where A6.2 binds, but the only escape mutations identified to date fall well outside the contact regions of both 2D3 and A6.2. To determine how mutations in distal residues could block antibody binding, we used molecular dynamics flexible fitting simulations of the atomic structures placed into the density map to examine the 2D3/MNV1 complex and these mutations. Our findings suggest that the escape mutant, V339I, may stabilize a salt bridge network at the P-domain dimer interface that, in an allostery-like manner, affects the conformational relaxation of the P domain and the efficiency of binding. They further highlight the unusual antigenic surface bound by MAb 2D3, one which elicits cross-reactive antibodies but which the virus is unable to alter to escape neutralization. These results may be leveraged to generate norovirus (NoV) vaccines containing broadly neutralizing antibodies. The simplest and most common way for viruses to escape antibody neutralization is by mutating residues that are essential for antibody binding. Escape mutations are strongly selected for by their effect on viral fitness, which is most often related to issues of protein folding, particle assembly, and capsid function. The studies presented here demonstrated that a broadly neutralizing antibody to mouse norovirus binds to an exposed surface but that the only escape mutants that arose were distal to the antibody binding surface. To understand this finding, we performed an analysis that suggested that those escape mutations blocked antibody binding by affecting structural plasticity. This kind of antigenic region-one that gives rise to broadly neutralizing antibodies but that the virus finds difficult to escape from-is therefore ideal for vaccine development.
理想的抗病毒疫苗能诱导产生具有广泛毒株识别能力的抗体（Abs），这些抗体可结合病毒难以通过突变来逃逸的区域。我们使用10种鼠诺如病毒（MNV）毒株和5种人诺如病毒（HuNoV）病毒样颗粒（VLPs），鉴定出了单克隆抗体（MAb）2D3，它能广泛中和所有测试的MNV毒株。重要的是，与该抗体对应的逃逸突变体产生得非常缓慢，且与针对我们之前研究的抗体A6.2产生的逃逸突变体位置不同。为了了解2D3中和作用的原子细节，我们确定了2D3/MNV1复合物的冷冻电子显微镜（cryo-EM）结构。有趣的是，2D3结合在P结构域的顶部，非常靠近A6.2的结合位点，但迄今为止鉴定出的唯一逃逸突变位于2D3和A6.2的接触区域之外。为了确定远端残基的突变如何阻断抗体结合，我们使用分子动力学柔性拟合模拟，将原子结构放入密度图中以研究2D3/MNV1复合物及这些突变。我们的研究结果表明，逃逸突变体V339I可能稳定了P结构域二聚体界面处的盐桥网络，该网络以类似变构的方式影响P结构域的构象松弛和结合效率。这些结果进一步凸显了MAb 2D3所结合的异常抗原表面，该表面能诱导产生交叉反应性抗体，但病毒无法通过改变该表面来逃逸中和作用。这些结果可用于开发含有广泛中和抗体的诺如病毒（NoV）疫苗。病毒逃逸抗体中和作用最简单、最常见的方式是突变抗体结合所必需的残基。逃逸突变因其对病毒适应性的影响而被强烈选择，这通常与蛋白质折叠、颗粒组装和衣壳功能等问题有关。此处展示的研究表明，一种针对小鼠诺如病毒的广泛中和抗体结合在一个暴露表面上，但产生的唯一逃逸突变体位于抗体结合表面之外。为了理解这一发现，我们进行了一项分析，结果表明那些逃逸突变通过影响结构可塑性来阻断抗体结合。因此，这种抗原区域——能产生广泛中和抗体但病毒难以逃逸的区域——对于疫苗开发来说是理想的。
