Catabolite Repression Regulation Research Articles

Development of an auto-inducible expression system by nitrogen sources switching based on the nitrogen catabolite repression regulation

BackgroundThe construction of protein expression systems is mainly focused on carbon catabolite repression and quorum-sensing systems. However, each of these regulatory modes has an inherent flaw, which is difficult to overcome. Organisms also prioritize using different nitrogen sources, which is called nitrogen catabolite repression. To date, few gene regulatory systems based on nitrogen catabolite repression have been reported.ResultsIn this study, we constructed a nitrogen switching auto-inducible expression system (NSAES) based on nitrogen catabolite regulation and nitrogen utilization in Aspergillus nidulans. The PniaD promoter that is highly induced by nitrate and inhibition by ammonia was used as the promoter. Glucuronidase was the reporter protein. Glucuronidase expression occurred after ammonium was consumed in an ammonium and nitrate compounding medium, achieving stage auto-switching for cell growth and gene expression. This system maintained a balance between cell growth and protein production to maximize stress products. Expressions of glycosylated and secretory proteins were successfully achieved using this auto-inducible system.ConclusionsWe described an efficient auto-inducible protein expression system based on nitrogen catabolite regulation. The system could be useful for protein production in the laboratory and industrial applications. Simultaneously, NSAES provides a new auto-inducible expression regulation mode for other filamentous fungi.

Transcriptional analysis of impacts of glycerol transporter 1 on methanol and glycerol metabolism in Pichia pastoris.

The efficient promoter of alcohol oxidase 1 (PAOX1) in methylotrophic yeast Pichia pastoris is strictly induced by methanol but repressed by glycerol with an unclear molecular mechanism. In the present study, the gene of a previously characterized transmembrane protein glycerol transporter 1 (GT1) of P. pastoris GS115 was deleted by homologous recombination. Transcriptional profiles of the mutant (gt1Δ) and wild type (WT) were compared with different carbon sources (glycerol, methanol and glycerol-methanol mix) at various time points using high-throughput RNA-Seq techniques. We determined that the loss of glycerol transporter 1 (Gt1p) could relieve catabolite repression in the glycerol-methanol mixed medium and shared a similar transcriptional profile with the WT in methanol medium. By calculating the common differentially expressed genes in three distinct paired groups, genes involved in the stress response, nutrition deprivation and translational process were identified, explaining the potential roles of glycerol in the regulation of methanol metabolism. Based on weighted gene co-expression network analysis, the relationship between biological traits and the transcriptional profile was established. With the support of published research and our data, we propose two possible regulatory pathways that are involved in the regulation of catabolite repression (adenosine 5΄-monophosphate (AMP)-activated protein kinase /SNF1 and Mitogen-activated protein kinase/HOG), thereby providing potential targets for both research and industrial strain improvement.

Beta-glucoside permease represses the bgl operon of Escherichia coli by phosphorylation of the antiterminator protein and also interacts with glucose-specific enzyme III, the key element in catabolite control.

The beta-glucoside (bgl) operon of Escherichia coli is subject to both positive control by transcriptional termination/antitermination and negative control by the beta-glucoside-specific transport protein, an integral membrane protein known as enzyme IIBgl. Previous results led us to speculate that enzyme IIBgl exerts its negative control by phosphorylating and thereby inactivating the antiterminator protein, BglG. Specifically, our model postulated that the transport protein enzyme IIBgl exhibits protein-phosphotransferase activity in the absence of beta-glucosides. We now present biochemical evidence that the phosphorylation of protein BglG does indeed occur in vivo and that it is accompanied by the loss of antitermination activity. BglG persists in the phosphorylated state in the absence of beta-glucosides but is rapidly dephosphorylated when beta-glucosides become available for transport. Our data also suggested specific interactions between the beta-glucoside transport protein and the glucose-specific enzyme III (enzyme IIIGlc), a component of glucose transport and a key element in regulation of catabolite repression. These observations indicate that enzyme IIIGlc may, in conjunction with enzyme IIBgl, modulate the transport of beta-glucosides and the phosphorylation of the antiterminator protein. In the absence of both sugars, when the catabolite-controlled promoter of the operon is derepressed, enzyme IIIGlc may mediate tight repression of antitermination.

Catabolite repression in Escherichia coli mutants lacking cyclic AMP

The regulation of catabolite repression of beta-galactosidase has been studied in Escherichia coli mutants deleted for the adenyl cyclase gene (cya delta), and thus unable to synthesize cyclic AMP. It has been found that, provided a second mutation occurs either in the crp gene coding for the catabolite gene activator protein (CAP) or in the Lactose region, these mutants exhibit catabolite repression. If the catabolite repression seen in the mutant strains corresponds to the mechanism operating in wild-type cells the results would suggest that the intracellular concentration of cyclic AMP cannot be the unique regulator of catabolite repression.

Catabolite modulator factor: Physiological properties and in vivo effects

Catabolite modulator factor (CMF) specifically inhibits the expression of operons sensitive to catabolite repression. Systems known to be catabolite independent are not affected by CMF. The rate of metabolism of CMF depends on the extent of catabolite repression: it is slow under conditions of strong repression and high in catabolically derepressed cells. Cyclic AMP does not interfere with the rate of CMF metabolism. It has been found that a certain class of crp mutants are partially resistant to the repressive effect of CMF. Our results provide considerable support for the existence of an additional negative control in the regulation of catabolite repression.