title: Direct measurements of IPTG enable analysis of the induction behavior of E. coli in high cell density cultures.
abstract: BACKGROUND
  The E. coli lac operon and its components have been studied for decades, and lac-derived systems are widely used for recombinant protein production. However, lac operon dynamics and induction behavior remain the paradigm of gene regulation. Recently, an HPLC-MS-based method to quantify IPTG in the medium and inside the biomass has been established, and this tool may be useful to uncover the lack of knowledge and allow optimization of biotechnological processes.
  
  
  RESULTS
  The results obtained from the study of IPTG distribution profiles in fed-batch, high cell density cultures allowed discrimination between two different depletion patterns of an inducer from the medium to the biomass in E. coli-expressing rhamnulose-1-phosphate aldolase (RhuA). Moreover, we could demonstrate that active transport mediates the uptake of this gratuitous inducer. Additionally, we could study the induction behaviors of this expression system by taking into account the biomass concentration at the induction time.
  
  
  CONCLUSIONS
  In the bistable range, partial induction occurred, which led to intermediate levels of RhuA activity. There was a direct relationship between the initial inducer concentrations and the initial inducer transport rate together with the specific activity. A majority of the inducer remains in the medium to reach equilibrium with the intracellular level. The intracellular inducer accumulation was a further evidence of bistability of the lac operon.
authors.affiliation: Departament d'Enginyeria Química, Unitat de Biocatàlisi Aplicada associada al IQAC (CSIC), Universitat Autònoma de Barcelona, Bellaterra, Spain.
authors.forename: Alfred
authors.initials: A
authors.lastname: Fernández-Castané
authors.forename: Glòria
authors.initials: G
authors.lastname: Caminal
authors.forename: Josep
authors.initials: J
authors.lastname: López-Santín
mesh_terms: D000446:Aldehyde-Lyases / Q000235:genetics / Q000378:metabolism; D018533:Biomass; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D007544:Isopropyl Thiogalactoside / Q000032:analysis* / Q000378:metabolism
chemical_list: D007544:Isopropyl Thiogalactoside; D000446:Aldehyde-Lyases; C022526:rhamnulose-1-phosphate adolase