Structural dynamics of the aggregation in globular proteins: an experimental study

The research activity during my PhD was dedicated to investigate the unfolding and aggregation properties of the globular proteins β-lactoglobulin (BLG) and bovine serum albumin (BSA), having in mind their potential use as building blocks to develop new functional materials. The thermal and pH-dependent aggregation mechanisms of BLG are characterized using a combination of linear and nonlinear spectroscopies. By focusing on both acidic and neutral pH conditions, distinct formations of molten globule states are revealed. Time-resolved techniques enhance the ability to deconvolve spectral features, providing deeper insights into the aggregation processes involved. In addition to the importance of aggregation, the formation of mixed materials is also explored due to their applications in various fields, such as food industry. The behavior of BLG-BSA mixtures is analyzed, leading to the successful creation of composite gels with a partial fibrillar nature at high concentrations. Additionally, dye chromophores are employed to exploit energy transfer processes related to protein aggregation. The characterization of the aggregation process exploiting FRET by labeling BLG with ad-hoc synthesized chromophores is achieved either through linear and time resolved spectroscopies. The obtained results lay the foundations for the potential application of various dye systems to characterize the interactions between proteins of different kind, extending the versatility of the methodology for studying co-aggregation in protein mixtures. This work provides valuable insights into the dynamics of protein aggregation, highlighting the role of specific molecular interactions and spectroscopic markers. It establishes a foundation for the future development of protein-based materials with controllable aggregation properties.