Influence of RNA Binding on the Structure and Dynamics of the Lassa Virus Nucleoprotein

Abstract

Lassa virus protects its viral genome through the formation of a ribonucleoprotein complex in which the nucleoprotein (NP) encapsidates the single-stranded RNA genome. Crystal structures provide evidence that a conformational change must occur to allow for RNA binding. In this study, the mechanism by which NP binds to RNA and how the conformational changes in NP are achieved was investigated with molecular-dynamics simulations. NP was structurally characterized in an open configuration when bound to RNA and in a closed form in the absence of RNA. Our results show that when NP is bound to RNA, the protein is highly dynamic and the system undergoes spontaneous deviations away from the open-state configuration. The equilibrium simulations are supported by free-energy calculations that quantify the influence of RNA on the free-energy surface, which governs NP dynamics. We predict that the globally stable states are qualitatively in agreement with the observed crystal structures, but that both open and closed conformations are thermally accessible in the presence of RNA. The free-energy calculations also provide a prediction of the location of the transition state for RNA binding and identify an intermediate metastable state that exhibits correlated motions that could promote RNA binding.

T‐705‐modified ssRNA in complex with Lassa virus nucleoprotein exhibits nucleotide splaying and increased water influx into the RNA‐binding pocket

“…The closed state may look ideal at first glance because it is the RNA-bound state but further investigation using MD simulation revealed that transition between open (b) and close occurs even in RNA-bound LASV-NP. Indeed, open-state RNA-bound LASV-NP did transition toward the closed LASV-NP structure as early as 120 ns (Pattis & May, 2016), and our group has independently confirmed this transition (data not shown) within the timescale of 200 ns in RNA-bound and Apo-NP and therefore started the simulations in the current study in the closed-state conformations in its Apo, unsubstituted ssRNA (ssRNA), single-T-705substituted (ssRNA+1), and four-T-705-substituted ssRNA (ssRNA+4) biosystems (Figure 1b).…”

“…The loops adjacent to α6 segment (aa: 113–127 and aa: 146–163) were also modeled using MODELER ( ver . 9.15) as previously described (Pattis & May, ). The complex with an unmodified ssRNA (5′‐UAUCUC‐3′) in complex with open LASV‐NP is called open‐RNA‐bound structure or unsubstituted biosystem.…”

Markov State Model of Lassa Virus Nucleoprotein Reveals Large Structural Changes during the Trimer to Monomer Transition

“…The dynamics of the N-terminal domain have been studied previously (Pattis and May, 2016). It was found that the slow global motions primarily involve helix 6 motions.…”

Genomic Diversity and Spatiotemporal Distributions of Lassa Virus Outbreaks in Nigeria

“…Out of all LASV proteins that exist in the virions and infected host cells, nucleoprotein has the largest number of polypeptide encapsidated genomic RNA to prevent it from degradation in infected host cells [10]. LASV nucleoprotein made up a peptide having 569 amino acids residue [11] and consists of a separate amine group (N-terminal) and carboxylic group (C-terminal) domains [12]. The N-terminal domain proposed a cap-binding cellular structure through deep binding cavity for the synthesis of viral mRNA [13], while the C-terminal domain revealed a binding site functions as exoribonuclease, leading to suppression of type I IFN production by interfering with IFN regulatory factor 3 (IRF-3)…”

“…activation [12,14,15]. This pathway plays an important role in transcription and replication of Lassa virus and immunosuppression of infected host [11,16] LASV glycoprotein (GP) is a trimeric single polypeptide chain, glycosylated cotranslationally, cleaved subunit GP-1 and GP-2 by enzyme peptidase in the endoplasmic reticulum (ER) which play an important role in viral-host infection [17,18] Glycoprotein facilitates the entry of the virion into the host cell through receptor binding and fusion to the cell membrane as the only unique protein of the capsid that protect humoral immunity [17]. Before the GP-1 attachment to the host cells, there is a low immune response that poses a major problem to the host immune system as the virus spike in response to protective antibodies.…”