Intrinsic Disorder and Salt-dependent Conformational Changes of the N-terminal TFIP11 Splicing Factor

Tuftelin Interacting Protein 11 (TFIP11) was recently identified as a critical human spliceosome assembly regulator. Indeed, it is involved in many biological functions including the interaction with multiple spliceosome key proteins and its localisation in several membrane-less organelles. However, there is a lack of structural information on TFIP11, which limits the rationalisation of its biological role. TFIP11 has been predicted as a highly disordered protein, and specifically concerning its N-terminal (N-TER) region. Intrinsically disordered proteins (IDPs) lack a defined tertiary structure, existing as a dynamic conformational ensemble, which favours their role as hubs in protein-protein and protein-RNA interaction networks. Furthermore, IDPs are involved in liquid-liquid phase separation (LLPS) which drives the formation of subnuclear compartments. In this study, we have refined the disorder prediction of TFIP11 N-TER region and subsequently performed all-atom molecular dynamics (MD) simulations to assess its conformational flexibility and the interplay between their different N-TER domains. We further confirm that TFIP11 may be described as a polyampholyte IDP with a flexible conformation. Furthermore, since LLPS formation and IDP conformational changes are salt-dependent phenomena we have investigated by MD simulations the influence of salt concentration in shaping the conformational ensemble of the N-TER region of TFIP11. Increasing the salt concentration enhances the flexibility of the TFIP11 N-TER conformation, which presents a fuzzier conformational landscape, a more globular shape, and an unstructured arrangement that could favor LLPS segregation and protein-RNA interaction.