title: Colloidal Stability &amp; Conformational Changes in β-Lactoglobulin: Unfolding to Self-Assembly.
abstract: A detailed understanding of the mechanism of unfolding, aggregation, and associated rheological changes is developed in this study for β-Lactoglobulin at different pH values through concomitant measurements utilizing dynamic light scattering (DLS), optical microrheology, Raman spectroscopy, and differential scanning calorimetry (DSC). The diffusion interaction parameter kD emerges as an accurate predictor of colloidal stability for this protein consistent with observed aggregation trends and rheology. Drastic aggregation and gelation were observed at pH 5.5. Under this condition, the protein's secondary and tertiary structures changed simultaneously. At higher pH (7.0 and 8.5), oligomerizaton with no gel formation occurred. For these solutions, tertiary structure and secondary structure transitions were sequential. The low frequency Raman data, which is a good indicator of hydrogen bonding and structuring in water, has been shown to exhibit a strong correlation with the rheological evolution with temperature. This study has, for the first time, demonstrated that this low frequency Raman data, in conjunction with the DSC endotherm, can be been utilized to deconvolve protein unfolding and aggregation/gelation. These findings can have important implications for the development of protein-based biotherapeutics, where the formulation viscosity, aggregation, and stability strongly affects efficacy or in foods where protein structuring is critical for functional and sensory performance.
authors.affiliation: Malvern Instruments, 7221 Lee Deforest Drive, Suite 300, Columbia, MD 21046, USA. steve.blake@malvern.com.
authors.forename: Steven
authors.initials: S
authors.lastname: Blake
authors.affiliation: Malvern Instruments, 7221 Lee Deforest Drive, Suite 300, Columbia, MD 21046, USA. samiul.amin@malvern.com.
authors.forename: Samiul
authors.initials: S
authors.lastname: Amin
authors.affiliation: Malvern Instruments, 7221 Lee Deforest Drive, Suite 300, Columbia, MD 21046, USA. wei.qi@malvern.com.
authors.forename: Wei
authors.initials: W
authors.lastname: Qi
authors.affiliation: Malvern Instruments, 7221 Lee Deforest Drive, Suite 300, Columbia, MD 21046, USA. madhabi.majumdar@malvern.com.
authors.forename: Madhabi
authors.initials: M
authors.lastname: Majumdar
authors.affiliation: Malvern Instruments, 7221 Lee Deforest Drive, Suite 300, Columbia, MD 21046, USA. neil.lewis@malvern.com.
authors.forename: E Neil
authors.initials: EN
authors.lastname: Lewis
mesh_terms: D003102:Colloids / Q000737:chemistry*; D007782:Lactoglobulins / Q000737:chemistry*; D011487:Protein Conformation; D055503:Protein Multimerization; D055550:Protein Stability; D058767:Protein Unfolding
chemical_list: D003102:Colloids; D007782:Lactoglobulins
keywords: Raman spectroscopy; differential scanning calorimetry; dynamic light scattering; microrheology; protein aggregation; protein unfolding; self-assembly