单股,阳性RNA((+)RNA)病毒在病毒诱导的胞内膜区室中复制其基因组。(+)RNA病毒通过编码膜相互作用蛋白和/或蛋白结构域,将它们的小基因组(几千到几万个碱基)的重要部分用于产生这些区室。诺罗病毒是(+)RNA病毒的非常多样化的属,包括人和动物病原体。人类诺如病毒是世界范围内急性胃肠炎的主要病因,基因型II基因型4(GII.4)诺如病毒占绝大多数感染。在病毒复制的N端编码的三种病毒蛋白多蛋白指导与诺如病毒复制相关的胞内膜重排。在这三个人中,非结构蛋白4(NS4)似乎是最重要的,尽管其在复制细胞器形成中的确切功能尚不清楚。在这里我们生产,纯化和表征GII.4NS4。AlphaFold建模与实验数据相结合,完善并校正了我们以前的NS4的粗略结构模型。使用简单的人工脂质体,我们报道了NS4膜特性的广泛表征。我们发现NS4自组装并由此将脂质体桥接在一起。Cryo-EM,NMR和膜浮选显示形成了几个不同的NS4组件,其中至少有两个以不同的方式将一对膜桥接在一起。诺罗病毒属于(+)RNA病毒,其复制区室从靶内膜挤出并产生双膜囊泡。我们的数据确定21kDaGII.4人诺如病毒NS4可以,在没有任何其他因素的情况下,在图波中概括了几个特征,包括膜并置,发生在这样的过程中。
Single-stranded, positive-sense RNA ((+)RNA) viruses replicate their genomes in virus-induced intracellular membrane compartments. (+)RNA viruses dedicate a significant part of their small genomes (a few thousands to a few tens of thousands of bases) to the generation of these compartments by encoding membrane-interacting proteins and/or protein domains. Noroviruses are a very diverse genus of (+)RNA viruses including human and animal pathogens. Human noroviruses are the major cause of acute gastroenteritis worldwide, with genogroup II genotype 4 (GII.4) noroviruses accounting for the vast majority of infections. Three viral proteins encoded in the N-terminus of the viral replication polyprotein direct intracellular membrane rearrangements associated with norovirus replication. Of these three, nonstructural protein 4 (NS4) seems to be the most important, although its exact functions in replication organelle formation are unknown. Here we produce, purify and characterize GII.4 NS4. AlphaFold modeling combined with experimental data refine and correct our previous crude structural model of NS4. Using simple artificial liposomes, we report an extensive characterization of the membrane properties of NS4. We find that NS4 self-assembles and thereby bridges liposomes together. Cryo-EM, NMR and membrane flotation show formation of several distinct NS4 assemblies, at least two of them bridging pairs of membranes together in different fashions. Noroviruses belong to (+)RNA viruses whose replication compartment is extruded from the target endomembrane and generates double-membrane vesicles. Our data establish that the 21-kDa GII.4 human norovirus NS4 can, in the absence of any other factor, recapitulate in tubo several features, including membrane apposition, that occur in such processes.