title: Increasing the efficiency of protein export in Escherichia coli.
abstract: Export of recombinant proteins to the periplasm of Escherichia coli is in many cases preferable to cytoplasmic production. However, when the protein is overexpressed, export efficiency decreases significantly and some advantages of the system are lost. This is what happens when attempting to produce recombinant human interleukin-6 (hIL-6) as a pre(OmpA) fusion in E. coli. Assuming that the host protein export machinery becomes overloaded, we have tested the effect of providing the host with additional copies of two key components of that machinery. Supplementation with a plasmid bearing prlA4 (secY allele) and secE genes increased the ratio of mature to precursor hIL-6 from 1.2 to 10.8. The increase in processing ratio was associated with the accumulation of a larger amount of total (mature plus precursor forms) hIL-6. Providing a plasmid-borne wild-type prlA was ineffective compared to prlA4 allele. This suggests that the PrlA protein, a component of the translocator, recognizes features at the mature portion of secretory substrates independently of those at the signal peptide portion.
authors.affiliation: Research Department, Antibióticos Farma S.A. Antonio López, Madrid, Spain.
authors.forename: J
authors.initials: J
authors.lastname: Pérez-Pérez
authors.forename: G
authors.initials: G
authors.lastname: Márquez
authors.forename: J L
authors.initials: JL
authors.lastname: Barbero
authors.forename: J
authors.initials: J
authors.lastname: Gutiérrez
mesh_terms: D001425:Bacterial Outer Membrane Proteins / Q000096:biosynthesis; D003001:Cloning, Molecular / Q000379:methods*; D004926:Escherichia coli / Q000378:metabolism*; D005798:Genes, Bacterial; D006801:Humans; D015850:Interleukin-6 / Q000096:biosynthesis*; D010957:Plasmids; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis*; D011994:Recombinant Proteins / Q000096:biosynthesis*
chemical_list: D001425:Bacterial Outer Membrane Proteins; D015850:Interleukin-6; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins