FC0066
Nucleoprotein Ntail (N)  -  Phosphoprotein (P)

Biological function
In paramyxoviruses, genome transcription and replication rely on the N-P interaction that is critical for the recruitment of the polymerase. This interaction makes the viral RNA accessible to the polymerase L through a structural reorganisation of the nucleocapsid.

Domain organization/sequence features
The HeV nucleoprotein (N) consists of two domains: a folded domain (referred to as N_CORE, residues 1–400) that is responsible for the interaction with the viral RNA and for maintaining the nucleocapsid structure, and an intrinsically disordered domain (referred to as N_TAIL, residues 401–532) that is responsible for the interaction with the P protein. RNA is accommodated in a positively charged pocket at the hinge between the two domains.

Structural evidence
XD interacts only locally with the MoRE(473-492) of N_TAIL and that no additional residues are involved in the interaction. The NMR signal intensities outside the MoRE are not perturbed by the presence of XD. The similarity of the high- resolution NMR spectroscopic parameters measured in the presence and absence of the partner, which together represent the signature that the N-terminal part of N_TAIL remains sterically hindered between turns of the nucleocapsid, indicates that there is no major reorganization of the nucleocapsid upon interaction. No new peaks appeared in the N_TAIL spectrum even with saturating amounts of P_XD. This suggests that even when bound to P_XD, HeV N_TAIL remains dynamic, undergoing exchange between different conformers on the P_XD surface. Despite the α-helical transition that HeV N_TAIL undergoes upon binding to P_XD, the experimentally determined RS of the NiV N_TAIL-P_XD complex suggests that binding to P_XD does not imply formation of a compact complex, instead it retains a considerable flexibility. In further support of this observation, the many observable and relatively sharp NMR resonances in HeV N_TAIL-P_XD complex, displaying chemical shifts that are nearly unaltered, provide evidence that N_TAIL remains significantly disordered even in the bound form. Therefore the final complex is likely endowed with flexible appendages in a structural arrangement possibly reminiscent of that observed in the case of the MeV complex.

Structure/Mechanism
High resolution NMR spectroscopy, as well as previous biochemical data, suggest that HeV N_TAIL-XD forms a so-called ’fuzzy’ complex. The complex between N_TAIL and XD appears to be dynamic, as the resonances of the MoRE of N_TAIL do not reappear even for large saturating amounts of XD, suggesting helical fraying of the MoRE of N_TAIL on the surface of XD. The interaction between XD and N_TAIL in HeV appears to be controlled by a combination of long-range electrostatic forces that correctly orient N_TAIL prior to accommodation in the narrow hydrophobic pocket on the surface of XD. The same intensities and chemical shifts, and therefore the same conformational signature, for these residues upon interaction with the X domain of the phosphoprotein provides evidence that the environment of N_TAIL is effectively identical in the free and XD-bound form of the nucleocapsids. Flanking fuzzy regions serve as a platform for the recruitment of additonal (regulatory) partners such as hsp70.

Mechanism category
Competitive binding

Significance
Fuzziness enables XD to be accommodated on N_TAIL without triggering a nucleocapsid rearrangement that could increase the accessibility of the polymerase complex to the viral genome.

Submitted by
Sonia Longhi   sonia.longhi@afmb.univ-mrs.fr