Heterologous protein export in Escherichia coli: influence of bacterial signal peptides on the export of human interleukin 1β

Expression plasmids carrying the coding sequence of mature human interleukin 1β(ILIβ) linked either to a Met start codon, or fused to different efficient Escherichia coli secretion signal sequences, have been constructed. In the latter case, we used signal peptides derived either from an outer membrane protein (OmpA) or from a periplasmic protein (PhoA). The synthesis of IL1β from these fusions was investigated in an otherwise strictly isogenic context using identical conditions of derepression and culture media. The Met-IL1β fusion produced a soluble cytoplasmic protein which could be released from the cells by osmotic shock whereas the OmpA and PhoA fusions were always insoluble. The extent of sOmpA-IL1βmaturation was found to vary from 50 to 100%, mainly depending on the medium used, whereas no significant maturation of the signal peptide could be detected in the case of the sPhoA-IL1β fusion. Immuno-electron microscopy revealed that the sOmpA-IL 1β fusion was targeted to the inner membrane, whereas the ThoA-IL 1βfusion remained within the cytoplasm and thus did not appear to enter the secretion pathway. Amplifying the E. coli signal peptidase lep gene on a multicopy plasmid did not improve signal peptide removal from sOmpA-IL1β. Moreover, these E. coli