Molecular characterization of Cowpea main storage protein breakdown products, generated during seed germination

Sprouting of legumes influence their nutritional quality: protein content increases, fat decreases, as well as antinutritional compounds [1]. Furthermore, as in case of most legumes, amino acid profile of cowpea complements cereal grain. Seed proteins are the nitrogen reserve that support seedling growth during first steps of germination. New findings indicate that several biological activities emerge after proteolytic breakdown, such as plant defense and antimicrobial properties [2], while bioactive peptide production, useful for human wellbeing, deserves investigations. Aim of this work is to study the proteolytic products originating from the major Cowpea storage protein β-vignin, after its treatment with proteases extracted from germinating cowpea seeds. This characterization was achieved by Thermal Shift Assay, under different conditions of pH and ionic strength. Secondly, the aggregation propensity was evaluated using a combination of chromatographic and electrophoretic techniques. The main finding is the molecular and structural characterization of 27 kDa intermediate breakdown polypeptides, highly resistant to proteolysis, that remain associated in a form similar to the native protein. It is thus likely that this core acts as a scaffold to the other proteolytic products that are however available to dissociate and to provide either nutrition to the germinating seed, or to exert some, still unexplored, bioactivity. It was recently described that pea seeds vicilin cupin domains can form digestion resistant amyloids [3], thus it is worth exploring this occurrence in the case of β-vignin as well, in order to elucidate the physiological function of the described breakdown products, their potential as bioactive peptides, and to further increase knowledge on plant amyloid biology, a still underexplored field. [1] Devi CB et al. (2015) J Food Sci Technol 52, 6821-7 [2] Jimenez-Lopez JC et al. (2016) Front. Plant Sci. 7, 1856 [3] Antonets KS et al. (2020) PLoS Biol 18(7): e3000564