title: Development of a highly efficient protein-secreting system in recombinant Lactobacillus casei.
abstract: The available techniques for heterologous protein secretion in Lactobacillus strains are limited. The aim of the present study was to develop an efficient protein-secretion system using recombinant lactobacilli for various applications such as live delivery of biotherapeutics. For the construction of expression vectors, the Lactobacillus brevis slpA promoter, Lactobacillus casei prtP signal sequence, and mouse IL-10 sequences were used as a model system. Interestingly, the slpA promoter exhibited strong activity in L. casei contrary to previous observations. In order to stabilize replication of the plasmid in E. coli, a removable terminator sequence was built into the promoter region. For the improvement of secretion efficiency, a DTNSD oligopeptide was added to the cleavage site of signal peptidase. The resulting plasmids provided remarkably efficient IL-10 secretion. Accumulation of the protein in the culture supernatant varied widely according to the pH conditions. By analysis of the secreted protein, formation of homodimers and biological activity, IL-10 was confirmed to be functional. The presently constructed plasmids could be useful tools for heterologous protein-secretion in L. casei.
authors.affiliation: Department of Food, Bioprocessing, and Nutrition Sciences, North Carolina State University, Box 7624, Raleigh, NC 27695, USA.
authors.forename: Akinobu
authors.initials: A
authors.lastname: Kajikawa
authors.forename: Eiko
authors.initials: E
authors.lastname: Ichikawa
authors.forename: Shizunobu
authors.initials: S
authors.lastname: Igimi
mesh_terms: D000818:Animals; D001483:Base Sequence; D005110:Extracellular Space / Q000235:genetics / Q000378:metabolism*; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism; D016753:Interleukin-10 / Q000235:genetics* / Q000378:metabolism*; D007780:Lacticaseibacillus casei / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D015202:Protein Engineering / Q000379:methods*; D021381:Protein Transport
chemical_list: D016753:Interleukin-10