Insights into the Hendra virus NTAIL–XD complex: Evidence for a parallel organization of the helical MoRE at the XD surface stabilized by a combination of hydrophobic and polar interactions

• The Hendra virus XD domain (XD) binds to the MoRE of NTAIL that spans aa 470–490.
• Upon binding, the MoRE folds into an α-helix and is embedded at the XD surface.
• The binding interface is hydrophobic, but the surrounding charged residues might play a role.
• Through mutational and binding studies we show that polar interactions do play a role.
• Mutational and SAXS studies support a parallel orientation of the MoRE with respect to XD.

The Hendra virus is a member of the Henipavirus genus within the Paramyxoviridae family. The nucleoprotein, which consists of a structured core and of a C-terminal intrinsically disordered domain (NTAIL), encapsidates the viral genome within a helical nucleocapsid. NTAIL partly protrudes from the surface of the nucleocapsid being thus capable of interacting with the C-terminal X domain (XD) of the viral phosphoprotein. Interaction with XD implies a molecular recognition element (MoRE) that is located within NTAIL residues 470–490, and that undergoes α-helical folding. The MoRE has been proposed to be embedded in the hydrophobic groove delimited by helices α2 and α3 of XD, although experimental data could not discriminate between a parallel and an antiparallel orientation of the MoRE. Previous studies also showed that if the binding interface is enriched in hydrophobic residues, charged residues located close to the interface might play a role in complex formation. Here, we targeted for site directed mutagenesis two acidic and two basic residues within XD and NTAIL. ITC studies showed that electrostatics plays a crucial role in complex formation and pointed a parallel orientation of the MoRE as more likely. Further support for a parallel orientation was afforded by SAXS studies that made use of two chimeric constructs in which XD and the MoRE were covalently linked to each other. Altogether, these studies unveiled the multiparametric nature of the interactions established within this complex and contribute to shed light onto the molecular features of protein interfaces involving intrinsically disordered regions.