title: An agmatine-inducible system for the expression of recombinant proteins in Enterococcus faecalis.
abstract: BACKGROUND
  Scientific interest in Enterococcus faecalis has increased greatly over recent decades. Some strains are involved in food fermentation and offer health benefits, whereas others are vancomycin-resistant and cause infections that are difficult to treat. The limited availability of vectors able to express cloned genes efficiently in E. faecalis has hindered biotechnological studies on the bacterium's regulatory and pathogenicity-related genes. The agmatine deiminase (AGDI) pathway of E. faecalis, involved in the conversion of agmatine into putrescine, is driven by a response inducer gene aguR.
  
  
  RESULTS
  This study describes that the exposure to the induction factor (agmatine) results in the transcription of genes under the control of the aguB promoter, including the aguBDAC operon. A novel E. faecalis expression vector, named pAGEnt, combining the aguR inducer gene and the aguB promoter followed by a cloning site and a stop codon was constructed. pAGEnt was designed for the overexpression and purification of a protein fused to a 10-amino-acid His-tag at the C-terminus. The use of GFP as a reporter of gene expression in E. faecalis revealed that under induction with 60 mM agmatine, fluorescence reached 40 arbitrary units compared to 0 in uninduced cells.
  
  
  CONCLUSIONS
  pAGEnt vector can be used for the overexpression of recombinant proteins under the induction of agmatine in E. faecalis, with a close correlation between agmatine concentration and fluorescence when GFP was used as reporter.
authors.affiliation: Instituto de Productos Lácteos de Asturias, IPLA-CSIC, Paseo Río Linares s/n, 33300, Villaviciosa, Asturias, Spain. daniml@ipla.csic.es.
authors.forename: Daniel M
authors.initials: DM
authors.lastname: Linares
authors.affiliation: Instituto de Productos Lácteos de Asturias, IPLA-CSIC, Paseo Río Linares s/n, 33300, Villaviciosa, Asturias, Spain. mpgarcia@ipla.csic.es.
authors.forename: Marta
authors.initials: M
authors.lastname: Perez
authors.affiliation: Instituto de Productos Lácteos de Asturias, IPLA-CSIC, Paseo Río Linares s/n, 33300, Villaviciosa, Asturias, Spain. ladero@ipla.csic.es.
authors.forename: Victor
authors.initials: V
authors.lastname: Ladero
authors.affiliation: Instituto de Productos Lácteos de Asturias, IPLA-CSIC, Paseo Río Linares s/n, 33300, Villaviciosa, Asturias, Spain. beadelrio@ipla.csic.es.
authors.forename: Beatriz
authors.initials: B
authors.lastname: Del Rio
authors.affiliation: Instituto de Productos Lácteos de Asturias, IPLA-CSIC, Paseo Río Linares s/n, 33300, Villaviciosa, Asturias, Spain. bredruello@ipla.csic.es.
authors.forename: Begoña
authors.initials: B
authors.lastname: Redruello
authors.affiliation: Instituto de Productos Lácteos de Asturias, IPLA-CSIC, Paseo Río Linares s/n, 33300, Villaviciosa, Asturias, Spain. mcm@ipla.csic.es.
authors.forename: M Cruz
authors.initials: MC
authors.lastname: Martin
authors.affiliation: Instituto de Productos Lácteos de Asturias, IPLA-CSIC, Paseo Río Linares s/n, 33300, Villaviciosa, Asturias, Spain. mfernandez@ipla.csic.es.
authors.forename: María
authors.initials: M
authors.lastname: Fernandez
authors.affiliation: Instituto de Productos Lácteos de Asturias, IPLA-CSIC, Paseo Río Linares s/n, 33300, Villaviciosa, Asturias, Spain. maag@ipla.csic.es.
authors.forename: Miguel A
authors.initials: MA
authors.lastname: Alvarez
mesh_terms: D000376:Agmatine / Q000494:pharmacology*; D013293:Enterococcus faecalis / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects*; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000235:genetics; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics
chemical_list: D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins; D000376:Agmatine