Data on the standardization of a cyclohexanone-responsive expression system for Gram-negative bacteria

Abstract

Engineering of robust microbial cell factories requires the use of dedicated genetic tools somewhat different from those traditionally used for laboratory-adapted microorganisms. We have edited and formatted the ChnR/PchnB regulatory node of Acinetobacter johnsonii to ease the targeted engineering of ectopic gene expression in Gram-negative bacteria. The proposed compositional standard was thoroughly verified with a monomeric and superfolder green fluorescent protein (msf•GFP) in Escherichia coli. The expression data presented reflect a tightly controlled transcription initiation signal in response to cyclohexanone. Data in this article are related to the research paper “Genetic programming of catalytic Pseudomonas putida biofilms for boosting biodegradation of haloalkanes” [1].

The Metabolic Redox Regime of Pseudomonas putida Tunes Its Evolvability toward Novel Xenobiotic Substrates

“…A synthetic metabolic module was designed for this purpose, in which the gene encoding NADH oxidase from S. pneumoniae was placed under control of the orthogonal ChnR/ P chnB expression system ( Fig. 4c ), which is inducible by cyclohexanone ( 74 , 75 ). We tested the effect of such an NADH-burning device on the overall physiology of P. putida EM⋅DNT by measuring the specific Nox activity in cell-free extracts and evaluating growth rates on succinate cultures ( Fig.…”

Improved Thermotolerance of Genome‐Reduced Pseudomonas putida EM42 Enables Effective Functioning of the PL/cI857 System

“…In order to benchmark the P L ‐ c I857 pair for operating in P. putida we adopted the SEVA standard for arraying regulatory elements in a genetic node shaped as a heterologous expression cargo. This standard, which was first adopted in the platform for the cyclohexanone‐responsive expression system of pSEVA2311 was implemented on the P L and c I857 parts as shown in Figure A and B. First, the c I857 mutant allele of the c I repressor of lambda phage was placed under the control of a constitutive promoter and a TIR motif upstream of the start codon.…”

Pyridine nucleotide transhydrogenases enable redox balance of Pseudomonas putida during biodegradation of aromatic compounds

“…We constructed another stress transcriptional reporter by amplifying the region −376 to −4 upstream of katB using oligonucleotides katB _TF‐F and katB _TF‐R (Table S2 in the Supporting Information). The resulting amplicon, spanning the promoter region of katB , was inserted into the SacI/BamHI sites of the pBBR1‐based broad‐range‐host vector pSEVA237M harbouring a promoterless msf · gfp gene (Silva‐Rocha et al ., ; Benedetti et al ., ). The plasmids containing the corresponding fusions (pSR· ahpC , pSR· katA and pSR· katB , Table S2 in the Supporting Information) were electroporated into P .…”