Dataset Information

Inducer exclusion, by itself, cannot account for the glucose-mediated lac repression of Escherichia coli.

ABSTRACT: The lac operon of Escherichia coli is repressed several 100-fold in the presence of glucose. This repression has been attributed to cAMP receptor protein-mediated inhibition of lac transcription and EIIA^Glc-mediated inhibition of lactose transport (inducer exclusion). The growing evidence against the first mechanism has led to the postulate that the repression is driven by inducer exclusion. Although inducer exclusion reduces the permease activity only 2-fold in fully induced cells, it could be more potent in partially induced cells. Here, we show that even in partially induced cells, inducer exclusion reduces the permease activity no more than 6-fold. Moreover, the repression is so small because these experiments are performed in the presence of chloramphenicol. Indeed, when glucose is added to a culture growing on glycerol and TMG, but no chloramphenicol, lac expression is repressed 900-fold. This repression is primarily due to reversal of the positive feedback loop, i.e., the decline of the intracellular TMG level leads to a lower permease level, which reduces the intracellular TMG level even further. The repression in the absence of chloramphenicol is therefore primarily due to positive feedback, which does not exist during measurements of inducer exclusion.

SUBMITTER: Aggarwal RK

PROVIDER: S-EPMC8943701 | biostudies-literature | 2022 Mar

REPOSITORIES: biostudies-literature

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Inducer exclusion, by itself, cannot account for the glucose-mediated lac repression of Escherichia coli.

Aggarwal Ritesh Kumar RK Narang Atul A

Biophysical journal 20220120 5

The lac operon of Escherichia coli is repressed several 100-fold in the presence of glucose. This repression has been attributed to cAMP receptor protein-mediated inhibition of lac transcription and EIIA<sup>Glc</sup>-mediated inhibition of lactose transport (inducer exclusion). The growing evidence against the first mechanism has led to the postulate that the repression is driven by inducer exclusion. Although inducer exclusion reduces the permease activity only 2-fold in fully induced cells, i ...[more]

PMID: 35065916

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Project description:As advances are made toward the industrial feasibility of mass-producing biofuels and commodity chemicals with sugar-fermenting microbes, high feedstock costs continue to inhibit commercial application. Hydrolyzed lignocellulosic biomass represents an ideal feedstock for these purposes as it is cheap and prevalent. However, many microbes, including Escherichia coli, struggle to efficiently utilize this mixture of hexose and pentose sugars due to the regulation of the carbon catabolite repression (CCR) system. CCR causes a sequential utilization of sugars, rather than simultaneous utilization, resulting in reduced carbon yield and complex process implications in fed-batch fermentation. A mutant of the gene encoding the cyclic AMP receptor protein, crp*, has been shown to disable CCR and improve the co-utilization of mixed sugar substrates. Here, we present the strain construction and characterization of a site-specific crp* chromosomal mutant in E. coli BL21 star (DE3). The crp* mutant strain demonstrates simultaneous consumption of glucose and xylose, suggesting a deregulated CCR system. The proteomics further showed that glucose was routed to the C5 carbon utilization pathways to support both de novo nucleotide synthesis and energy production in the crp* mutant strain. Metabolite analyses further show that overflow metabolism contributes to the slower growth in the crp* mutant. This highly characterized strain can be particularly beneficial for chemical production by simultaneously utilizing both C5 and C6 substrates from lignocellulosic biomass.IMPORTANCEAs the need for renewable biofuel and biochemical production processes continues to grow, there is an associated need for microbial technology capable of utilizing cheap, widely available, and renewable carbon substrates. This work details the construction and characterization of the first B-lineage Escherichia coli strain with mutated cyclic AMP receptor protein, Crp*, which deregulates the carbon catabolite repression (CCR) system and enables the co-utilization of multiple sugar sources in the growth medium. In this study, we focus our analysis on glucose and xylose utilization as these two sugars are the primary components in lignocellulosic biomass hydrolysate, a promising renewable carbon feedstock for industrial bioprocesses. This strain is valuable to the field as it enables the use of mixed sugar sources in traditional fed-batch based approaches, whereas the wild-type carbon catabolite repression system leads to biphasic growth and possible buildup of non-preferential sugars, reducing process efficiency at scale.

Dataset Information

Inducer exclusion, by itself, cannot account for the glucose-mediated lac repression of Escherichia coli.

Publications

Inducer exclusion, by itself, cannot account for the glucose-mediated lac repression of Escherichia coli.

Similar Datasets

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