Intrinsically disordered proteins (IDPs) display a large number of interaction modes including folding-upon-binding, binding without major structural transitions, or binding through highly dynamic, so-called fuzzy, complexes. The vast majority of experimental information about IDP binding modes have been inferred from crystal structures of proteins in complex with short peptides of IDPs. However, crystal structures provide a mainly static view of the complexes and do not give information about the conformational dynamics experienced by the IDP in the bound state. Knowledge of the dynamics of IDP complexes is of fundamental importance to understand how IDPs engage in highly specific interactions without concomitantly high binding affinity. Here, we combine rotating-frame R, Carr-Purcell-Meiboom Gill relaxation dispersion as well as chemical exchange saturation transfer to decipher the dynamic interaction profile of an IDP in complex with its partner. We apply the approach to the dynamic signaling complex formed between the mitogen-activated protein kinase (MAPK) p38α and the intrinsically disordered regulatory domain of the MAPK kinase MKK4. Our study demonstrates that MKK4 employs a subtle combination of interaction modes in order to bind to p38α, leading to a complex displaying significantly different dynamics across the bound regions.

The genome of influenza A virus (IAV) comprises eight RNA segments (vRNA) which are transcribed and replicated by the heterotrimeric IAV RNA-dependent RNA-polymerase (RdRp). RdRp consists of three subunits (PA, PB1 and PB2) and binds both the highly conserved 3′- and 5′-ends of the vRNA segment. The IAV RdRp is an important antiviral target, but its structural mechanism has remained largely elusive to date. By applying a polyprotein strategy, we produced RdRp complexes and define a minimal human IAV RdRp core complex. We show that PA-PB1 forms a stable heterodimeric submodule that can strongly interact with 5′-vRNA. In contrast, 3′-vRNA recognition critically depends on the PB2 N-terminal domain. Moreover, we demonstrate that PA-PB1 forms a stable and stoichiometric complex with host nuclear import factor RanBP5 that can be modelled using SAXS and we show that the PA-PB1-RanPB5 complex is no longer capable of 5′-vRNA binding. Our results provide further evidence for a step-wise assembly of IAV structural components, regulated by nuclear transport mechanisms and host factor binding.