In-Silico Analysis of pH-Dependent Liquid-Liquid Phase Separation in Intrinsically Disordered Proteins

This work was funded by European Union's Horizon 2020 research and innovation programme under GA 952334 (PhasAGE). S.V. was supported by the Spanish Ministry of Science and Innovation (PID 2019-105017RB-I00) and by ICREA Academia 2020. C.P.-G. was supported by the Secretariat of Universities and Research of the Catalan Government and the European Social Fund (2021 FI_B 00087). Intrinsically disordered proteins (IDPs) are essential players in the assembly of biomolecular condensates during liquid-liquid phase separation (LLPS). Disordered regions (IDRs) are significantly exposed to the solvent and, therefore, highly influenced by fluctuations in the microenvironment. Extrinsic factors, such as pH, modify the solubility and disorder state of IDPs, which in turn may impact the formation of liquid condensates. However, little attention has been paid to how the solution pH influences LLPS, despite knowing that this process is context-dependent. Here, we have conducted a large-scale in-silico analysis of pH-dependent solubility and disorder in IDRs known to be involved in LLPS (LLPS-DRs). We found that LLPS-DRs present maximum solubility around physiological pH, where LLPS often occurs, and identified significant differences in solubility and disorder between proteins that can phase-separate by themselves or those that require a partner. We also analyzed the effect of mutations in the resulting solubility profiles of LLPS-DRs and discussed how, as a general trend, LLPS-DRs display physicochemical properties that permit their LLPS at physiologically relevant pHs.