Assessing induced folding within the intrinsically disordered C-terminal domain of the Henipavirus nucleoproteins by site directed spin labeling EPR spectroscopy
Abstract
This work aims at characterizing structural transitions within the intrinsically disordered C-terminal domain of the nucleoprotein (N TAIL) from the Nipah and Hendra viruses, two recently emerged pathogens gathered within the Henipavirus genus. To this end, we used site directed spin labeling combined with EPR spectroscopy to investigate the α-helical induced folding that Henipavirus N TAIL domains undergo in the presence of the C-terminal X domain of the phosphoprotein (P XD). For each N TAIL protein, six positions located within four previously proposed molecular recognition elements (MoREs) were targeted for spin labeling, with three of these (positions 475, 481, 487) falling within the MoRE responsible for binding to P XD (Box3). A detailed analysis of the impact of the partner protein on the labeled N TAIL variants revealed a dramatic modification in the environment of the spin labels grafted within Box3, with the observed modifications supporting the formation of an induced α-helix within this region. In the free state, the slightly lower mobility of the spin labels grafted within Box3 as compared to the other positions suggests the existence of a transiently populated α-helix, as already reported for measles virus (MeV) N TAIL. Comparison with the well-characterized MeV N TAIL-P XD system, allowed us to validate the structural models of Henipavirus N TAIL-P XD complexes that we previously proposed. In addition, this study highlighted a few notable differences between the Nipah and Hendra viruses. In particular, the observation of composite spectra for the free form of the Nipah virus N TAIL variants spin labeled in Box3 supports conformational heterogeneity of this partly pre-configured αhelix, with the pre-existence of stable α-helical segments. Altogether these results provide insights into the molecular mechanisms of the Henipavirus N TAIL-P XD binding reaction. Supplementary Material Supplementary material dealing with Figures S1 to S5 can be directly downloaded from the JBSD site.
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