Structure Determination of Feline Calicivirus Virus-Like Particles in the Context of a Pseudo-Octahedral Arrangement

The vesivirus feline calicivirus (FCV) is a positive strand RNA virus encapsidated by an icosahedral T=3 shell formed by the viral VP1 protein. Upon its expression in the insect cell - baculovirus system in the context of vaccine development, two types of virus-like particles (VLPs) were formed, a majority built of 60 subunits (T=1) and a minority probably built of 180 subunits (T=3). The structure of the small particles was determined by x-ray crystallography at 0.8 nm resolution helped by cryo-electron microscopy in order to understand their formation. Cubic crystals belonged to space group P213. Their self-rotation function showed the presence of an octahedral pseudo-symmetry similar to the one described previously by Agerbandje and co-workers for human parvovirus VLPs. The crystal structure could be solved starting from the published VP1 structure in the context of the T=3 viral capsid. In contrast to viral capsids, where the capsomers are interlocked by the exchange of the N-terminal arm (NTA) domain, this domain is disordered in the T=1 capsid of the VLPs. Furthermore it is prone to proteolytic cleavage. The relative orientation of P (protrusion) and S (shell) domains is alerted so as to fit VP1 to the smaller T=1 particle whereas the intermolecular contacts around 2-fold, 3-fold and 5-fold axes are conserved. By consequence the surface of the VLP is very similar compared to the viral capsid and suggests a similar antigenicity. The knowledge of the structure of the VLPs will help to improve their stability, in respect to a use for vaccination.猫杯状病毒（vesivirus feline calicivirus, FCV）是一种正链RNA病毒，由病毒VP1蛋白构成的二十面体T=3衣壳包裹。在疫苗研发背景下，于昆虫细胞-杆状病毒表达系统中表达该病毒蛋白时，可形成两种类型的病毒样颗粒（virus-like particles, VLPs）：多数由60个亚基组装而成（对应T=1型），少数则由180个亚基组装而成（对应T=3型）。为阐明该小颗粒的形成机制，研究人员结合冷冻电子显微镜（cryo-electron microscopy）辅助手段，通过X射线晶体学以0.8 nm的分辨率解析了其结构。该立方晶体的空间群为P213。其自身旋转函数（self-rotation function）分析显示存在八面体伪对称性，这与此前Agerbandje及其团队针对人细小病毒（human parvovirus）VLPs所报道的伪对称性一致。该晶体结构可基于已发表的T=3型病毒衣壳VP1结构进行解析求解。与天然病毒衣壳不同，天然病毒衣壳的衣壳亚基通过N端臂（N-terminal arm, NTA）结构域的交换实现互锁；而该VLPs的T=1型衣壳中，NTA结构域处于无序状态，且易于被蛋白酶水解。此外，为使VP1适配更小的T=1型颗粒，P（突出）结构域（protrusion domain, P domain）与S（壳体）结构域（shell domain, S domain）的相对取向发生了改变，而围绕2次、3次和5次对称轴的分子间相互作用仍得以保留。因此，该VLPs的表面与天然病毒衣壳极为相似，提示二者具有相似的抗原性。对该VLPs结构的解析将有助于优化其稳定性，以推动其在疫苗接种中的应用。