In silico Analysis of Different Signal Peptides for the Excretory Production of Recombinant NS3-GP96 Fusion Protein in Escherichia coli

Escherichia coli is one of the simplest hosts which is widely being used to express heterologous proteins. However, without appropriate signal peptide, this host cannot be applied for secretory proteins. Secretory production of recombinant proteins in E. coli has been an issue of interest because of its diverse advantages including cost and time savings, as well as reduction of endotoxin. NS3 from hepatitis C virus (HCV) was chosen as an antigen for vaccine development against HCV virus infections and it connected to gp96 as an adjuvant for stimulating Toll-like receptors (TLRs) to stimulate cytokines secretion by T cells. It was successfully produced in E. coli without using signal peptide previously. In this study, in order to increase the expression level of recombinant NS3-gp96 fusion protein (rNS3-gp96) in periplasmic space, we selected a series of signal peptides. Therefore, to foretell the best signal peptides for expression of NS3-gp96 recombinant protein in E. coli, 52 signal peptides from gram-negative bacteria were chosen and the most important physicochemical features of them were investigated. Therefore, n, h and c regions and signal peptide probability of them were evaluated by signalP software “version 4.1”, and physicochemical features were assessed by ProtParam and PROSO II tools. Eventually, prsK protein, outer membrane pore protein E (phoE), and fimbrial adapter papK protein were determined as the best candidates for the secretory production of rNS3-gp96 in E. coli in our study (with D score 0.899, 0.806, 0.797, respectively). Although, in the experimental investigation, should be considered other influencing parameters.