Macromolecular Crowding Promotes Reentrant Liquid-Liquid Phase Separation of Human Serum Transferrin and Prevents Surface-Induced Fibrillation

Abstract

Protein aggregation and inactivation upon surface immobilization are major limiting factors for analytical applications in biotechnology related fields. Protein immobilization on solid surfaces often requires multi‐step surface passivation which is time consuming and inefficient. Herein, we have discovered that biomolecular condensates of biologically active human serum transferrin (Tf) can effectively prevent surface‐induced fibrillation and preserve the native‐like conformation of phase separated Tf over a period of 30‐days. It has been observed that macromolecular crowding promotes homotypic liquid‐liquid phase separation (LLPS) of Tf through enthalpically driven multivalent hydrophobic interactions possibly via the involvement of its low complexity domain (residue 3–20) containing hydrophobic amino acids. The present LLPS of Tf is a rare example of salt-mediated reentrant phase separation in a broad range of salt concentrations (0–3 M) solely via the involvement of hydrophobic interactions.