title: Protein expression vector and secretion signal peptide optimization to drive the production, secretion, and functional expression of the bacteriocin enterocin A in lactic acid bacteria.
abstract: Replacement of the leader sequence (LS) of the bacteriocin enterocin A (LS(entA)) by the signal peptides (SP) of the protein Usp45 (SP(usp45)), and the bacteriocins enterocin P (SP(entP)), and hiracin JM79 (SP(hirJM79)) permits the production, secretion, and functional expression of EntA by different lactic acid bacteria (LAB). Chimeric genes encoding the SP(usp45), the SP(entP), and the SP(hirJM79) fused to mature EntA plus the EntA immunity genes (entA+entiA) were cloned into the expression vectors pNZ8048 and pMSP3545, under control of the inducible P(nisA) promoter, and in pMG36c, under control of the constitutive P(32) promoter. The amount, antimicrobial activity, and specific antimicrobial activity of the EntA produced by the recombinant Lactococcus lactis, Enterococcus faecium, E. faecalis, Lactobacillus sakei and Pediococcus acidilactici hosts varied depending on the signal peptide, the expression vector, and the host strain. However, the antimicrobial activity and the specific antimicrobial activity of the EntA produced by most of the LAB transformants was lower than expected from their production. The supernatants of the recombinant L. lactis NZ9000 (pNZUAI) and L. lactis NZ9000 (pNZHAI), overproducers of EntA, showed a 1.2- to 5.1-fold higher antimicrobial activity than that of the natural producer E. faecium T136 against different Listeria spp.
authors.affiliation: Departamento de Nutrición, Bromatología y Tecnología de los Alimentos, Facultad de Veterinaria, Universidad Complutense de Madrid, 28040 Madrid, Spain.
authors.forename: Juan
authors.initials: J
authors.lastname: Borrero
authors.forename: Juan J
authors.initials: JJ
authors.lastname: Jiménez
authors.forename: Loreto
authors.initials: L
authors.lastname: Gútiez
authors.forename: Carmen
authors.initials: C
authors.lastname: Herranz
authors.forename: Luis M
authors.initials: LM
authors.lastname: Cintas
authors.forename: Pablo E
authors.initials: PE
authors.lastname: Hernández
mesh_terms: D000900:Anti-Bacterial Agents / Q000378:metabolism* / Q000494:pharmacology; D001430:Bacteriocins / Q000096:biosynthesis* / Q000235:genetics / Q000494:pharmacology; D003001:Cloning, Molecular; D056546:Lactobacillales / Q000235:genetics / Q000378:metabolism*; D008087:Listeria / Q000187:drug effects; D013058:Mass Spectrometry; D008826:Microbial Sensitivity Tests; D021382:Protein Sorting Signals / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000494:pharmacology
chemical_list: D000900:Anti-Bacterial Agents; D001430:Bacteriocins; D021382:Protein Sorting Signals; D011994:Recombinant Proteins; C107799:enterocin A