Mechanistic exploration of Perfluorooctanoic Acid-Induced Hen egg white lysozyme aggregation at physiological Conditions: Spectroscopic and thermodynamic insights

Perfluoroalkyl substances (PFASs) have garnered attention in various scientific fields for their harmful effects on biological systems, including the endocrine, immune, and nervous systems. While research has delved into PFASprotein binding, their impact on protein aggregation remains uncharted. In this study, we investigated the effect of perfluorooctanoic acid (PFOA) on the aggregation of Hen Egg White Lysozyme (HEWL), a model protein known to aggregate under harsh conditions. We uncovered compelling insights by employing an array of spectroscopic, computational and thermal techniques. Isothermal titration calorimetry (ITC), molecular docking, and fluorescence spectroscopy reveal strong PFOA-HEWL interactions via hydrogen bonding, hydrophobic forces, and ionic interactions. Moreover, techniques like synchronous fluorescence spectroscopy, circular dichroism, and FTIR demonstrates that PFOA induces substantial conformational changes in HEWL, profoundly affecting its secondary structure. Increased PFOA concentrations have been observed to lead to HEWL aggregation under physiological conditions (pH 7.4 and room temperature), verified through various assays and imaging techniques. This is the first comprehensive exploration of PFOA's influence on protein aggregation. Our findings provide valuable insights into the health risks of PFOA exposure from a protein aggregation perspective, advancing our knowledge in environmental ecology, toxicology, pathology, and life sciences.