Sequence-Dependent Material Properties of Biomolecular Condensates and their Relation to Dilute Phase Conformations

Abstract

Material properties of phase-separated biomolecular assemblies, enriched with disordered proteins, dictate their ability to participate in many cellular functions. Despite the significant effort dedicated to understanding how the sequence of the disordered protein drives its phase separation to form condensates, little is known about the sequence determinants of condensate material properties. Here, we decipher these relationships for charged disordered proteins using model sequences comprised of glutamic acid and lysine residues as well as naturally occurring sequences of LAF1's RGG domain and DDX4's N-terminal domain. We do so by delineating how the arrangement of oppositely charged residues within these sequences influences the dynamical, rheological, and interfacial properties of the condensed phase. Interestingly, we find that the material properties of both the model and natural proteins respond similarly to the segregation of charges, despite their very different sequence compositions. Condensates of the highly charge-segregated sequences exhibit slower dynamics than the uniformly charge-patterned sequences, because of their comparatively long-lived molecular contacts between oppositely charged residues. Surprisingly, the molecular interactions within the condensate are highly similar to those within a single-chain for all sequences. Consequently, the condensate material properties of charged disordered proteins are strongly correlated with their dense-phase contact dynamics and their single-chain structural properties. Our findings demonstrate the potential to harness the sequence characteristics of disordered proteins for engineering the material properties of functional condensates, with insights from the dilute-phase properties.

Dissecting neurofilament tail sequence-phosphorylation-structure relationships with multicomponent reconstituted protein brushes

“…In a single-component brush, more evenly shuffling the NFM sequence charges caused the brush to expand further. This finding in the context of surface-tethered IDPs complements evidence from IDPs free in solution 32,34–36 and even specifically NFM tail fragments in solution 37 , where sequence charge desegregation tends to cause chain expansion. We observe that the charge shuffle effect on NFM tail brush height is negligible at low ionic strengths, possibly due to the lack of screening of significant charge repulsion between distant residues in the net-negative NFM sequence.…”

Determinants of Disordered Protein Co-Assembly Into Discrete Condensed Phases

“…This would suggest that FUS LC wildtype and LAF-1 RGG interactions are weakly attractive under experimental conditions, as EI close to 1 is observed, unlike partitioning of synthetic polymers such as PEG in aqueous two-phase systems in which EI << 1 63, 64 . Elevated concentrations of B at the interface for non-zero λ AB reflect locally enhanced concentration of client molecules due to favorable heterotypic interactions, which are not sufficiently strong to cause co-mixing but may be sufficient to alter condensates’ material properties and composition control via a thin interfacial layer 65–68 .…”

Conformational Properties of Polymers at Droplet Interfaces as Model Systems for Disordered Proteins

“…Intrinsically disordered proteins (IDPs) play an important role in the formation of biomolecular condensates through LLPS [6][7][8][9][10][11] . Due to the numerous similarities between IDPs and synthetic polymers, classical polymer-based models offer a powerful approach for investigating the conformations 12 , dynamics 13 , and phase behavior of IDPs [14][15][16][17] . In particular, such models have been extensively used to reveal the sequence-dependent conformations of IDPs in the dense and dilute phases of condensates [18][19][20][21][22][23][24][25][26][27] .…”

“…Lipiński et al reported that the condensate interface can serve as a nucleation site promoting the aggregation of amyloidogenic proteins 31 . Inspired by the critical role of interfaces in various functionalities, much research has focused on determining mesoscopic interfacial properties such as surface tension 13,32,33 , surface adsorption 34 , and electrochemical properties 35 , which are deemed important for regulating biological functions 29,36 . For example, it was shown that the condensates could form an autophagosome only if the surface tension is lower than a critical value 37 .…”