Thermodynamic stability of cowpea β-vignin: a calorimetric study to explore the impact of seed germination on protein native structure

β-vignin belongs to the 7S globulin or vicilin-like family and is the most represented storage protein in cowpea seeds (Vigna unguiculata, L. Walp) [1]. Recently, storage proteins have been shown not to be only a mere nitrogen reserve for seedling growth during germination, but new findings have revealed they are object of a selective breakdown of specific peptide bonds. Because of such regulated proteolytic events, some transient intermediate peptides are formed and have been shown to possess specific bioactivities [2]. β-vignin consists of two main differently glycosylated isoforms. Moreover, it exists in a monomer-trimer equilibrium that is affected by the environment pH: the higher the pH value, the greater the trimer-to-monomer ratio [3]. The full understanding of cowpea β-vignin physicochemical properties is crucial to reveal possible biological roles and to approach applicative uses in various fields, including eco-friendly plant defense, nutrition and nutraceutics. In this context, the main focus of this study was to explore the thermodynamic stability of both the cowpea native β-vignin and the structures produced during seed germination. To this aim, purified β-vignin was subjected to controlled proteolysis using cowpea proteases obtained from germinating seeds, mimicking the early germination process [3]. Subsequently, a calorimetric investigation was conducted by applying high-sensitivity DSC on both undigested and digested β-vignin under various buffered environments (pH 9.0, 8.5, and 6.5). Additionally, a thermodynamic analysis was performed on the undigested form of β-vignin to evaluate the protein stability and the thermal denaturation mechanism. Our findings revealed a distinct behavior of the storage protein compared to proteins serving other biological functions.