In-silico selection of appropriate signal peptides for romiplostim secretory production in Escherichia coli

Thrombocytopenia is a prevalent condition caused by a reduction in platelet production, increased platelet depletion, or a combination of both processes. Romiplostim, the most potent drug and the only thrombopoietin (TPO) receptor agonist, consists of four TPO receptor–activating peptides fused as a fusion protein to the human IgG1 heavy chain segment. Romiplostim could be produced by recombinant DNA technology in Escherichia coli. Extracellular and periplasmic secretion using signal peptides (SPs) of the recombinant proteins could have several advantages in purification and protein recovery. SPs are short amino-acid sequences used in protein translocation through various membranes. The structure of the different SPs identified in various bacteria and their compatibility with the protein could affect the transfer success and efficacy. This study aimed to identify the best SP for extracellular secretion of romiplostim to facilitate its purification and reduce production costs in E. coli. The amino acid sequences of the SPs and the position of their cleavage sites in the proteins were evaluated using the Signal 5.0 server. The solubility of the proteins produced in E. coli was predicted by protein-sol. The physicochemical parameters of the SPs were analyzed by ProtParam using the ExPASy server. We found that SfmC is the most suitable candidate, followed by TorT and bla. However, this result should be confirmed by further experimental assessment. Furthermore, for the successful secretion of a recombinant protein, the optimal growth conditions should be controlled in addition to a suitable SP.