Biophysical insights into pH-induced structural dynamics of recombinant interferon β-1b: Aggregation under acidic...

Recombinant interferon beta-1b (rIFN β-1b) is a therapeutic protein used to treat multiple sclerosis (MS). This study investigates the influence of solution conditions on the structural conformation of rIFN β-1b for better pharmaceutical formulations. The structural properties of rIFN β were studied at various pH values (2.0-12.0) using multiple biophysical techniques. Far UV CD and FTIR analysis confirmed the presence of α-helices in the native confirmation of rIFN β-1b as well as in alkaline conditions, exclusively. pH-dependent exposure of the tryptophan residues was probed by neutral (acrylamide) and ionic (iodide) quenching studies. At pH 5.0-12.0, rIFNβ-1b formed collisional complex with tryptophan at the excited state (dynamic quenching). Dye-binding studies validated the tertiary structural measurements by binding to the exposed hydrophobic patches and revealed protein aggregation between pH 2.0-4.0. This was further corroborated by dynamic light scattering (DLS) measurements. Differential scanning calorimetry (DSC) showed the thermodynamic measurements at various pHs. This investigation discusses about the comprehensive behaviour of rIFNβ-1b at various pH and establishes a foundation for advancements in pharmaceutical formulations and development.