Interaction-Dependent\nSecondary Structure of Peptides\nin Biomolecular Condensates

Biomolecular condensates provide a mechanism for compartmentalization\nof biomolecules in eukaryotic cells. These liquid-like condensates\nare formed via liquid–liquid phase separation, by a plethora\nof interactions, and can mediate several biological processes in healthy\ncells. Expansions of dipeptide repeat proteins, DPRs, in which arginine\nrich DPRs like poly-proline-arginine (PR), and poly-glycine-arginine\n(GR), partition RNA into condensates can however induce cell toxicity.\nHere, we use (GR)<sub>20</sub> as a model for biological poly-GR and\ncondense it using either excluded volume interactions with polyethylene\nglycol (PEG) as a crowder or direct electrostatic interactions with\nRNA oligomers. Using two-dimensional infrared (2D IR) spectroscopy,\nwe observe that (GR)<sub>20</sub> condensed through an excluded volume\nforms β-sheet structures, whereas (GR)<sub>20</sub> condensed\nwith RNA forms loops. We also investigate local hydrogen-bond dynamics\nin the condensate and compare the measurements with molecular dynamics\nsimulations. Hydrogen bond lifetimes undergo a marked slowdown compared\nto dynamics in the dilute phase. This is representative of confined\nwater within the percolated networks inside the condensate due to\nthe interaction present in the condensate disrupting H-bond networks.\nOverall, our results show that both protein structure and dynamics\nare inherently dependent on the type of interactions that stabilize\nthe condensates.