Post-Translational Modifications in Liquid-Liquid Phase Separation

Liquid-liquid phase separation (LLPS) is a phase change that is important in living organisms and is a unique way to form biomolecular condensates. When a cell releases a recruitment signal at a specific range and time, the original dispersed protein must undergo certain changes in order to "aggregate". After performing its function, the protein accepts the new signal and returns to solution without altering the existing intracellular protein expression level. Therefore, rapid protein post-translational modification (PTM) is an ideal way to regulate phase separation. PTM can modify the structure, charge, hydrophobicity and other properties of the proteins involved in phase separation, thus affecting phase transition behavior. The study of LLPS is still in its infancy, and the regulation of LLPS by PTM will undoubtedly be the focus of future research.

Fig. 1. Phosphorylation in LLPS. (Li J, et al., 2022)

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PTM can change the spatial structural properties, charge state and volume of the modified amino acids, thus affecting the activity state, localization, flip-flop and protein-protein interactions of the whole protein. Therefore, how PTM affects LLPS-induced bioaggregates is of great interest to us. Here at CD BioSciences, we can help our customers analyze the regulation of LLPS by various PTMs, including but not limited to:

• Phosphorylation
  Phosphorylation is one of the most common and important PTMs. We can derive the regulation of LLPS by phosphorylation in vitro by calculating the average charge maps of phosphorylated fragile X mental retirement protein (FMRP) and Tau protein.
• Arginine Methylation
  We co-express the proteins arginine methyltransferase and dead box polypeptide 4 (Ddx4) in E. coli to cause Ddx4N1 to be methylated at several sites, and further analyze the effect of arginine methylation on phase separation ability.
• Arginine Citrullination
  We can analyze the effect of citrullination catalyzed by peptidylarginine deiminase on protein aggregation, protein-protein interactions and protein phase separation.
• Acetylation
  We can analyze the effect of partial acetylation of dead cassette RNA decapping enzyme 3,X-linked (DDX3X) by lysine acetyltransferase binding protein (Ac-CBP) on its in vitro occurrence of LLPS.
• Ubiquitination
  We analyze the multivalent interactions of the structural domains of Ubiquilins and the effect on LLPS by NMR.
• Poly(ADP-ribosyl)-ation (PAR)
  We can analyze the effect of PAR-mediated protein-protein interactions on protein phase behavior.

RNA-binding proteins also undergo regulated phase transitions in a range of cell types. We also work to analyze the role of PTM in regulating RNA-binding protein condensation.

We can help you systematically understand the relationship between post-translational protein modifications and phase separation, and focus more on the changes in physical properties induced by the modifications in regulating phase separation. If you are interested in our services, please do not hesitate to contact us for more information.