Vennema H, Godeke G J, Rossen J W, Voorhout W F, Horzinek M C, Opstelten D J, Rottier P J
Department of Functional Morphology, Faculty of Veterinary Medicine, Utrecht University, The Netherlands.
EMBO J. 1996 Apr 15;15(8):2020-8. doi: 10.1002/j.1460-2075.1996.tb00553.x.
Budding of enveloped viruses has been shown to be driven by interactions between a nucleocapsid and a proteolipid membrane. By contrast, we here describe the assembly of viral envelopes independent of a nucleocapsid. Membrane particles containing coronaviral envelope proteins were assembled in and released from animal cells co-expressing these proteins' genes from transfected plasmids. Of the three viral membrane proteins only two were required for particle formation, the membrane glycoprotein (M) and the small envelope protein (E). The spike (S) protein was dispensable but was incorporated when present. Importantly, the nucleocapsid protein (N) was neither required not taken into the particles when present. The E protein, recently recognized to be a structural protein, was shown to be an integral membrane protein. The envelope vesicles were found by immunogold labelling and electron microscopy to form a homogeneous population of spherical particles indistinguishable from authentic coronavirions in size (approximately 100 nm in diameter) and shape. They were less dense than virions and sedimented slightly slower than virions in sucrose velocity gradients. The nucleocapsid-independent formation of apparently bona fide viral envelopes represents a novel mode of virus assembly.
有包膜病毒的出芽已被证明是由核衣壳与蛋白脂质膜之间的相互作用驱动的。相比之下，我们在此描述了独立于核衣壳的病毒包膜组装过程。含有冠状病毒包膜蛋白的膜颗粒在共表达转染质粒中这些蛋白基因的动物细胞内组装并释放。在三种病毒膜蛋白中，只有两种是颗粒形成所必需的，即膜糖蛋白（M）和小包膜蛋白（E）。刺突（S）蛋白并非必需，但存在时会被整合进去。重要的是，核衣壳蛋白（N）既不是必需的，存在时也不会被纳入颗粒中。最近被确认为结构蛋白的E蛋白被证明是一种整合膜蛋白。通过免疫金标记和电子显微镜发现，包膜囊泡形成了一群均匀的球形颗粒，其大小（直径约100nm）和形状与真正的冠状病毒难以区分。它们的密度比病毒粒子低，在蔗糖速度梯度中的沉降速度比病毒粒子稍慢。明显真实的病毒包膜的核衣壳非依赖性形成代表了一种新的病毒组装模式。
