In vitro assembly of Ebola virus nucleocapsidlike complex expressed in E. coil

Ebola virus （EBOV） harbors an RNA genome encapsi- dated by nucleoprotein （NP） along with other viral pro- teins to form a nucleocapsid complex. Previous Cryo- eletron tomography and biochemical studies have shown the helical structure of EBOV nucleocapsid at nanometer resolution and the first 450 amino-acid of NP （NP△451-739） alone is capable of forming a helical nucleocapsid-Iike complex （NLC）. However, the struc- tural basis for NP-NP interaction and the dynamic pro- cedure of the nucleocapsid assembly is yet poorly understood. In this work, we, by using an E. coli expression system, captured a series of images of NP△451-739 conformers at different stages of NLC assembly by negative-stain electron microscopy, which allowed us to picture the dynamic procedure of EBOV nucleocapsid assembly. Along with further biochemical studies, we showed the assembly of NLC is salt-sensi- tive, and also established an indispensible role of RNA in this process. We propose the diverse modes of NLC elongation might be the key determinants shaping the plasticity of EBOV virions. Our findings provide a new model for characterizing the self-oligomerization of viral nucleoproteins and studying the dynamic assembly process of viral nucleocapsid in vitro.