The CreC Regulator of Escherichia coli, a New Target for Metabolic Manipulations

Abstract

The CreBC (carbon source-responsive) two-component regulation system of Escherichia coli affects a number of functions, including intermediary carbon catabolism. The impacts of different creC mutations (a ⌬creC mutant and a mutant carrying the constitutive creC510 allele) on bacterial physiology were analyzed in glucose cultures under three oxygen availability conditions. Differences in the amounts of extracellular metabolites produced were observed in the null mutant compared to the wild-type strain and the mutant carrying creC510 and shown to be affected by oxygen availability. The ⌬creC strain secreted more formate, succinate, and acetate but less lactate under low aeration. These metabolic changes were associated with differences in AckA and LdhA activities, both of which were affected by CreC. Measurement of the NAD(P)H/NAD(P) ؉ ratios showed that the creC510 strain had a more reduced intracellular redox state, while the opposite was observed for the ⌬creC mutant, particularly under intermediate oxygen availability conditions, indicating that CreC affects redox balance. The null mutant formed more succinate than the wild-type strain under both low aeration and no aeration. Overexpression of the genes encoding phosphoenolpyruvate carboxylase from E. coli and a NADH-forming formate dehydrogenase from Candida boidinii in the ⌬creC mutant further increased the yield of succinate on glucose. Interestingly, the elimination of ackA and adhE did not significantly improve the production of succinate. The diverse metabolic effects of this regulator on the central biochemical network of E. coli make it a good candidate for metabolic-engineering manipulations to enhance the formation of bioproducts, such as succinate.T he survival of an organism depends, at least in part, on its ability to sense and respond to changes in the environment. In bacteria, global regulators control the transcription of genes in response to specific external stimuli and metabolic signals, finely tuning different aspects of their physiology to overcome environmental challenges. In Escherichia coli, seven global regulators (ArcA, Crp, Fis, Fnr, Ihf, Lrp, and NarL) directly modulate the expression of about one-half of all genes (1). This facultative aerobe is able to adapt its metabolism to different oxygen availability conditions through the concerted actions of a network of regulators, including the global regulators ArcAB and Fnr (2-4). These regulators affect many metabolic pathways, allowing the cells to reach an adequate redox balance under any given conditions. There is a very close association between carbon and electron flows, and even small differences in oxygen availability have been observed to elicit profound effects on the distribution of carbon fluxes (5).CreBC (for carbon source responsive) is a global sensing and regulation system affecting genes involved in a variety of functions, including enzymes of intermediary catabolism (6). Previous studies have shown that the creABCD operon is activated (i) during growth in minimal mediu...

A High-Throughput Method for Identifying Novel Genes That Influence Metabolic Pathways Reveals New Iron and Heme Regulation in Pseudomonas aeruginosa

“…MochiView-generated diagrams of Met-Seq enrichments of key genes mentioned in the text. (A) Genes or operons of (i) the creBC two-component signaling system ( 77 ), (ii) the vanAB vanillate synthesis operon, (iii) the phnAB anthranilate (PQS) biosynthesis operon, (iv) the tseF T6SS effector, (v) the fptA siderophore transporter gene, and (vi) the iron/siderophore associated ECF sigma factor PA1363. Note that neither fptA or tseF Tn insertion enrichments were included in Table S2 , as they did not meet the minimal read cutoff.…”

“…One example is creB ( PA0463 ) and creC ( PA0464 ) ( Table S1 ; Fig. S5A ), a two-component signaling system that regulates production of the TCA cycle entry molecule acetyl-CoA ( 77 ). Interestingly, the CreC histidine kinase recognizes a peptide derived from the iron-binding host protein lactoferrin, which is an important source of iron during infection ( 78 ).…”

Comparative genomic analysis of the Hafnia genus reveals an explicit evolutionary relationship between the species alvei and paralvei and provides insights into pathogenicity

“…MetD (MetI-N-Q) is a high-affinity transport system for methionine, which is an important amino acid involved in numerous metabolic processes in bacteria [13]. The CreC-CreB (carbon source-responsive) two-component regulation system in Escherichia coli affects a number of functions, including the intermediary carbon catabolism and intracellular redox state [14]. 2-O-α-mannosyl-D-glycerate is taken up by the MngA phosphotransferase system and utilized as a sole carbon source [15].…”

Refactoring the Embden–Meyerhof–Parnas Pathway as a Whole of Portable GlucoBricks for Implantation of Glycolytic Modules in Gram-Negative Bacteria

“… 69 The detailed protocol for the construction of this Δ glk mutant is described in the Supporting Information . Mutations in glycolytic genes were accumulated in E. coli derivatives via sequential transduction of individual alleles with bacteriophage P1 38 , 70 using individual mutants from the KEIO collection 71 as donors, followed by elimination of the antibiotic resistance marker using plasmid pCP20. 72 …”