title: Engineering of the gut commensal bacterium Bacteroides ovatus to produce and secrete biologically active murine interleukin-2 in response to xylan.
abstract: OBJECTIVE
  The aim of this work was to engineer a gut commensal bacterium, Bacteroidesovatus, to produce and secrete a biologically active cytokine in a regulated manner as a basis for novel immunotherapies for chronic gut disorders.
  
  
  RESULTS
  Bacteroides ovatus was engineered to produce murine interleukin-2 (MuIL2) intracellularly in response to xylan in culture media by inserting the MuIL2 gene into the xylanase operon of the organism. A second strain was engineered to secrete MuIL2 by adding Bacteroides fragilis enterotoxin secretion signal sequence to the protein. The recombinant strains produced MuIL2 only in the presence of xylan as determined by ELISA of cell lysates and culture supernatants. The IL2-dependent cell line CTLL-2 was used to demonstrate that MuIL2 produced by both B. ovatus strains was biologically active. This activity could be blocked by an anti-IL2 neutralizing antibody. The xylan-inducible nature of this system was demonstrated by RT-PCR.
  
  
  CONCLUSIONS
  Bacteroides ovatus was successfully engineered to produce and secrete biologically active MuIL2 in a xylan-inducible manner.
  
  
  CONCLUSIONS
  The production and secretion of a biologically active mammalian protein by a member of the gut microflora could lead to the development of new long-term immunotherapies for inflammatory gut diseases.
authors.affiliation: School of Biochemistry and Microbiology, University of Leeds, Leeds, UK.
authors.forename: M D
authors.initials: MD
authors.lastname: Farrar
authors.forename: T R
authors.initials: TR
authors.lastname: Whitehead
authors.forename: J
authors.initials: J
authors.lastname: Lan
authors.forename: P
authors.initials: P
authors.lastname: Dilger
authors.forename: R
authors.initials: R
authors.lastname: Thorpe
authors.forename: K T
authors.initials: KT
authors.lastname: Holland
authors.forename: S R
authors.initials: SR
authors.lastname: Carding
mesh_terms: D000818:Animals; D001439:Bacteroides / Q000235:genetics* / Q000276:immunology*; D019149:Bioreactors; D002460:Cell Line; D005818:Genetic Engineering; D006801:Humans; D007167:Immunotherapy; D007376:Interleukin-2 / Q000032:analysis / Q000096:biosynthesis* / Q000235:genetics; D007410:Intestinal Diseases / Q000628:therapy; D051379:Mice; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014990:Xylans / Q000494:pharmacology*
chemical_list: D007376:Interleukin-2; D014990:Xylans