Computational Design for the Production of Secretory Recombinant Codon-Optimized Human Stem Cell Factor (SCF) in Chinese Hamster Ovary (CHO) Cells With an Appropriate Signal Peptide: An Intensive In Silico Study

Background: Production of the recombinant proteins in mammalian cells is an important issue witha bio-therapeutic purpose. Numerous efforts have been focused on the improvement of the yields ofrecombinant proteins, which include optimization of conventional biological processes, selection ofappropriate signal peptides, codon optimization, and re-engineering of cells to produce more proteins.Stem cell factor (SCF) is a blood cytokine which activates the c-Kit receptor. This factor is crucial notonly for the differentiation of hematopoietic progenitor cells but also for the survival, proliferation, anddifferentiation of mast cells. Recently, its therapeutic role in several diseases such as Alzheimer’s andmyocardial infarction has been investigated. Therefore, the aim of this study was to design a secretoryrecombinant human SCF with the maximal yield in an appropriate mammalian host cell as Chinesehamster ovary (CHO) cells using the computational studies.Methods: As the first step, computational simulation studies were carried out to design the appropriatesignal peptide for the human SCF protein. Codon optimized coding sequence of hSCF was transferredinto a eukaryotic expression vector (pBudCE۴.۱). Recombinant vector (pBudCE۴.۱/SCF) was transfectedinto CHO cells and the stably transformed cells were screened and isolated. Subsequently, the expressionof SCF was defined by reverse transcription polymerase chain reaction (RT-qPCR) in stably transformedcells.Results: Our bioinformatics studies indicated that Azurocidin signal peptide could be a suitable signalpeptide for the production of SCF proteins in the CHO cells. Accordingly, computational studies revealedthat the presence of ۶×His-tag did not have a significant impact on the three-dimensional structure of theprotein. Furthermore, the expression of hSCF was significant in the stable CHO cells.Conclusion: The use of this approach may, therefore, lead to the production of highly efficient recombinanthSCF, which would be feasible for the mass production of this factor for therapeutic purposes.

کلیدواژه ها:

Chinese hamster ovary cells ، Codon optimization ، Signal peptide ، Stem cell factor