Probing protein structure in the context of biomolecular condensation

Molecular compartmentalisation is a central pillar of life. Spontaneous demixing of proteins and nucleic acids from solution via liquid-liquid phase separation has been identified as a key mechanism underlying the formation of dynamic membraneless compartments in the cell. Biomolecular condensates facilitate concentration of specific molecules at specific sites in response to specific environmental conditions.<br/>RNA-binding proteins are over-represented as drivers of condensate formation, and are often composed of both ordered (stable 3-dimensional fold) domains and intrinsically disordered (sample many conformations) regions (IDRs). Stable folded structures of proteins are generally well understood and are confidently predicted from their amino acid sequence. However, relationships between IDR composition, structure, material state and function are poorly understood. <br/>Using small-angle scattering techniques, X-ray crystallography, in vitro biophysical and cell-based assays, we aim to characterise both folded domains and IDRs to describe composition-structure-material relationships. Our model system is a family of abundant RNA-binding proteins in mammalian cells (DBHS proteins) that form condensates for regulating transcription, RNA-processing and DNA repair. DBHS proteins have a conserved core folded dimerisation domain, with extended intrinsically disordered regions at each terminus that are diverse in length and amino acid composition. We show that the IDRs of these proteins are central to their ability to undergo liquid demixing by phase separation in vitro and in the cell nucleus. We hypothesise that differential dimerisation of DBHS partners provides an avenue for tuning condensate formation and material properties in the cell, and provides a unique opportunity for investigating IDR composition-function relationships.