Utilizing immobilized recombinant serine alkaline protease from Bacillus safensis lab418 in wound healing: Gene cloning, heterologous expression, optimization, and characterization

• Published in: International journal of biological macromolecules Vol. 270; no. Pt 1; p. 132286
• Main Authors: El-Sayed, Ghada M., Agwa, Mona M., Emam, Maha T.H., Kandil, Heba, Abdelhamid, Ahmed E., Nour, Shaimaa A.
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.06.2024
• Subjects: Heterologous expression; Immobilization; Optimization; Serine alkaline protease; Wound healing; Immobilization; Wound healing; Serine alkaline protease; Optimization; Heterologous expression
• ISSN: 0141-8130; 1879-0003
• DOI: 10.1016/j.ijbiomac.2024.132286

Microbial proteases have proven their efficiency in various industrial applications; however, their application in accelerating the wound healing process has been inconsistent in previous studies. In this study, heterologous expression was used to obtain an over-yielding of the serine alkaline protease. The serine protease-encoding gene aprE was isolated from Bacillus safensis lab 418 and expressed in E. coli BL21 (DE3) using the pET28a (+) expression vector. The gene sequence was assigned the accession number OP610065 in the NCBI GenBank. The open reading frame of the recombinant protease (aprEsaf) was 383 amino acids, with a molecular weight of 35 kDa. The yield of aprEsaf increased to 300 U/mL compared with the native serine protease (SAFWD), with a maximum yield of 77.43 U/mL after optimization conditions. aprEsaf was immobilized on modified amine-functionalized films (MAFs). By comparing the biochemical characteristics of immobilized and free recombinant enzymes, the former exhibited distinctive biochemical characteristics: improved thermostability, alkaline stability over a wider pH range, and efficient reusability. The immobilized serine protease was effectively utilized to expedite wound healing. In conclusion, our study demonstrates the suitability of the immobilized recombinant serine protease for wound healing, suggesting that it is a viable alternative therapeutic agent for wound management. •The serine-alkaline protease gene (aprE) was properly expressed in E. coli.•An increase in activity of the recombinant protease (aprEsaf) – 9.3-fold as more as native enzyme.•The recombinant aprEsaf was successfully immobilized on modified amine-functionalized films (MAFs).•The immobilized enzyme exhibited distinctive thermal stability and efficient reusability at a wider pH range.•The final form of immobilized enzyme proved its efficiency in accelerating the process of wound healing.