title: Design and selection of a synthetic operon.
abstract: Cell-free systems are showing increasing promise for biosynthesis of both proteins and small molecules. However, in vitro transcription and translation reactions have so far primarily been used for the production of single proteins. In order to demonstrate the possibilities for coupled reactions, we designed synthetic operons that included different combinations of wild-type or evolved biotin ligases and streptavidins and demonstrated a mechanism for self-selection of operons following expression in vitro. Peptide substrates for biotin ligase were conjugated to the DNA operons and could be modified by a biotin ligase specific for either biotin or desthiobiotin and subsequently captured via a streptavidin specific for either biotin or desthiobiotin.
authors.affiliation: Institute of Cellular and Molecular Biology University of Texas at Austin , Austin, Texas, United States.
authors.forename: Wei-Cheng
authors.initials: WC
authors.lastname: Lu
authors.forename: Andrew D
authors.initials: AD
authors.lastname: Ellington
mesh_terms: D001710:Biotin / Q000031:analogs & derivatives / Q000138:chemical synthesis / Q000235:genetics; D002474:Cell-Free System / Q000737:chemistry; D004247:DNA / Q000737:chemistry / Q000235:genetics; D005786:Gene Expression Regulation; D053263:Gene Regulatory Networks; D005813:Genes, Synthetic; D009876:Operon / Q000235:genetics*; D012097:Repressor Proteins / Q000138:chemical synthesis / Q000235:genetics; D019809:Streptavidin / Q000138:chemical synthesis / Q000235:genetics; D058615:Synthetic Biology; D014158:Transcription, Genetic; D014177:Translations
chemical_list: D012097:Repressor Proteins; D001710:Biotin; C004749:desthiobiotin; D004247:DNA; D019809:Streptavidin