Role of Trifluoroacetic Acid Anions in pHP1α Liquid-Liquid Phase Separations

Heterochromatin Protein 1α (HP1α) is essential for chromatin organization and gene silencing, with its ability to undergo liquid-liquid phase separation (LLPS) playing a key role in heterochromatin maintenance. This process is influenced by phosphorylation of its N-terminal extension (NTE), electrostatic interactions with DNA, and binding of ligands at the chromo shadow domain (CSD). These multivalent interactions stabilize phase-separated droplets, which are critical for chromatin compaction and transcriptional regulation. While inorganic salts such as KCl modulate LLPS in vitro, they do not fully replicate cellular conditions where organic anions dominate. Here, we investigate potassium trifluoroacetate (CF₃COOK) as a substitute for potassium acetate in probing HP1α phase separation using solid-state NMR spectroscopy. CF₃COOK offers unique insights due to its distinct ¹⁹F signals. Comparing CF₃COOK and KCl, we explore ionic effects on HP1α dynamics and structure, revealing how these environments impact phase-separated states and advancing our understanding of chromatin-associated protein coacervation.