Ecombinant production and characterization of an okara-derived bioactive peptide

Bioactive peptides obtained from agricultural byproducts, such as soybean okara, represent a promising source of high-value molecules for sustainable applications [1]. However, selective isolation represents the major bottleneck, as direct purification from complex protein hydrolysates is highly inefficient, and traditional chemical synthesis is limited by high costs and environmental impact. To overcome these limitations, this study presents a sustainable and scalable biotechnological platform to produce a potential bioactive peptide identified from a screening of peptides originated from okara protein digestion [1]. Instead of chemical synthesis, the target peptide was produced using the recombinant IMPACT® (Intein-Mediated Purification with an Affinity Chitin-binding Tag) system in Escherichia coli BL21. The fusion protein was purified in a single step using chitin-affinity chromatography, exploiting the dithiothreitol (DTT)-induced self-cleaving activity of the intein tag to release the target peptide. This approach aims to achieve high-level expression and cost-effective purification, offering a green alternative to traditional chemical peptide synthesis. Successful soluble recombinant protein overexpression was achieved when the peptide was cloned at the N-terminal portion of the fusion protein. The peptide was then isolated, and residual DTT-required for intein self-cleavage was removed, yielding the pure target peptide, as confirmed by electrophoretic and chromatographic analyses. Moreover, peptide identity was by mass spectrometry analysis. Functional and structural characterization were performed to confirm the stability and integrity of the recombinant peptide. Its potential bioactivity in promoting plant growth and inhibiting fungal pathogens was also investigated. In conclusion, this work demonstrates the viability of combining bio-inspired sequence discovery from agricultural waste with green recombinant technologies for the sustainable production of functional peptides. References: [1] De Benedetti S, Girlando V, Pasquali M, Scarafoni A. Valorization of okara by enzymatic production of anti-fungal compounds for plant protection. Molecules. 2021; 26: 4858.