Investigation of secreted human fibroblast growth factor (hbFGF) production in the Bacillus subtilis

Recombinant protein production, as an advanced technology in biotechnology, plays a vital role in life sciences and medicine. Growth factors such as bFGF, due to their key role in wound healing and cell proliferation processes, are of great importance in the development of new therapies and in improving the quality of life of patients. The aim of this study was to produce recombinant human fibroblast growth factor (hbFGF) in the Bacillus subtilis strain to provide an effective and economical method for the production of this vital protein. In this research, the hbFGF gene was initially cloned into the PHT-43 vector, which was then transformed into E.coli. Following this, a secondary transformation was performed into Bacillus subtilis. Subsequently, the processes of expression, extraction, and purification of the protein were carried out. Finally, its biological activity was evaluated using assays on the NIH/3T3 cell line. The results demonstrated the successful expression and secretion of hbFGF protein in the Bacillus subtilis strain; however, the quantity of purified protein was not optimal compared to the nickel-column bound protein. Analyses indicated that various proteases in Bacillus subtilis contributed to the cleavage of part of the protein before the His-tag sequence, leading to a decrease in the purity of the final product. To address this issue, it is recommended to utilize engineered strains with greater efficiency that contain protease inhibitors, preventing the cleavage of the produced protein.