lac operon induction in Escherichia coli: Systematic comparison of IPTG and TMG induction and influence of the transacetylase LacA.

Titlelac operon induction in Escherichia coli: Systematic comparison of IPTG and TMG induction and influence of the transacetylase LacA.
Publication TypeJournal Article
Year of Publication2012
AuthorsMarbach, A, Bettenbrock, K
JournalJ Biotechnol
Volume157
Issue1
Pagination82-8
Date Published2012 Jan
ISSN1873-4863
KeywordsAcetyltransferases, beta-Galactosidase, Cell Count, Escherichia coli, Escherichia coli Proteins, Green Fluorescent Proteins, Isopropyl Thiogalactoside, Lac Operon, Methylgalactosides, Microscopy, Fluorescence, Monosaccharide Transport Proteins, Protein Engineering, Recombinant Proteins, Symporters, Thiogalactosides
Abstract

Most commonly used expression systems in bacteria are based on the Escherichia coli lac promoter. Furthermore, lac operon elements are used today in systems and synthetic biology. In the majority of the cases the gratuitous inducers IPTG or TMG are used. Here we report a systematic comparison of lac promoter induction by TMG and IPTG which focuses on the aspects inducer uptake, population heterogeneity and a potential influence of the transacetylase, LacA. We provide induction curves in E. coli LJ110 and in isogenic lacY and lacA mutant strains and we show that both inducers are substrates of the lactose permease at low inducer concentrations but can also enter cells independently of lactose permease if present at higher concentrations. Using a gfp reporter strain we compared TMG and IPTG induction at single cell level and showed that bimodal induction with IPTG occurred at approximately ten-fold lower concentrations than with TMG. Furthermore, we observed that lac operon induction is influenced by the transacetylase, LacA. By comparing two Plac-gfp reporter strains with and without a lacA deletion we could show that in the lacA(+) strain the fluorescence level decreased after few hours while the fluorescence further increased in the lacA(-) strain. The results indicate that through the activity of LacA the IPTG concentration can be reduced below an inducing threshold concentration-an influence that should be considered if low inducer amounts are used.

DOI10.1016/j.jbiotec.2011.10.009
Alternate JournalJ. Biotechnol.