Jennifer Tersteegen, Antti Mäkynen, Adam Harmat, Maria Sammalkorpi, Markus B. Linder, Jaakko V. I. Timonen
Liquid-liquid phase separation (LLPS) of biomolecules is key to biological processes and biomaterial formation (1). Since living systems operate under non-equilibrium conditions, factors like concentration and pH shifts impact LLPS. Understanding this dynamic behavior is essential to grasp cellular functions and biomaterial assembly.
Here, we study experimentally an engineered spider silk protein in vitro that undergoes LLPS and investigate its non-equilibrium behavior under the influence of an external electric field. The studied multidomain protein features an intrinsically disordered middle block, flanked by globular terminal domains. Brightfield microscopy and digital holographic microscopy reveal diverse, voltage-dependent behavior, including condensate movement, interfacial changes, reversible dissolution of condensates, and electro-convective patterns. We associate this behavior with electric field-driven pH changes due to electrolysis of water as well as electrophoresis and electrodiffusiophoresis. The pH changes affect, e.g., the charges of protein terminal domains – our previous work indicates that the interaction of the terminal domains is essential for the studied protein's ability to undergo LLPS (2). Understanding the far-from-equilibrium behavior of protein condensates can facilitate actively driven and controlled materials assembly.