Computational Investigation into the Solution Structure of Monomeric Lassa Virus Nucleoprotein and Insights into its RNA Binding Mechanism

Abstract

Lassa is a dangerous Arenavirus, which is enveloped and has a single-stranded (ss)RNA genome. The nucleoprotein (NP) is the most abundant protein in the virion and functions to encapsidate the viral genome to form a ribonucleoprotein complex (RNP). Studies of the full-length NP show it forms a circular trimer and does not bind to RNA. However, a structure of just the N-terminal domain bound to RNA has also been determined, which indicates structural reorganization is required to bind ssRNA. Therefor it is believed that NP must undergo a large conformational change from the observed trimeric configuration in order to bind and polymerize on ssRNA to form the RNP.Using molecular dynamics and metadynamics simulations we have investigated the influence of RNA on the global motions of the N-terminal domain of Lassa NP, and have identified that RNA-binding reduces the free energy barrier to transition to the RNA bound state. We have gone on to investigate the conformational dynamics of the full-length NP in apo and RNA bound states when free in solution as a monomer. During multi-microsecond MD simulations performed on the Anton supercomputer, we observed significant conformational changes in the NP. In both apo and RNA bound simulations, we observe changes in interdomain contacts and relative domain orientation, as well formation of helical secondary structure in an unresolved domain linker loop. Most notably in the RNA bound simulation, the domain orientation is much more dramatic, and provides a putative template to understand the higher-order structural organization of the RNP. Kinetic network analysis and Brownian dynamics simulations have been employed to further analyze the structural transition pathways and to evaluate mechanisms of transition from the RNA-free trimer to the RNP.