Reducing the glucose uptake rate in <i>Escherichia coli</i> affects growth rate but not protein production

Picón, Antonia; Mattos, M. Joost Teixeira de; Postma, Pieter W.

doi:10.1002/bit.20387

Cited by 40 publications

(35 citation statements)

References 39 publications

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“…In the literature, ptsG mutant strains have been reported to exhibit 20-to 40%-reduced growth rates compared to those of the parent strains in minimal media (4,5,44,45). Picon et al (44,45) described a ptsG manXYZ double mutant that exhibited a growth rate of 0.13 h Ϫ1 (19% of that of the parent strain), which is significantly higher than that observed in our experiments (Fig. 1).…”

Section: Discussioncontrasting

confidence: 62%

“…This could be due to the low glucose uptake rates in these mutants avoiding overflow metabolism (45). Acetate production still occurred in some ptsG mutant strains described in the literature (4,45), probably due to a different strain background (MC4100). In the abovementioned fast-growing ptsG strains, acetate excretion was observed, but the acetate yield was low.…”

Section: Discussionmentioning

confidence: 97%

“…Mutational analysis under aerobic and oxygen-limited conditions revealed an enrichment of ptsG mutations in anaerobic cultures, in contrast to mutations for enhanced mgl expression aerobically (35), indicating the different importances of these systems depending on oxygen availability. Mutant strains defective in the glucose PTS (deletion of ptsG, ptsH, ptsI, or crr) have been previously investigated (4,5,17,26,39,45). These mutants have a reduced growth rate in glucose minimal medium and exhibit a strongly reduced rate of acetate formation (45) even in complex media, where the growth rate is unaffected (4).…”

mentioning

confidence: 99%

“…Mutant strains defective in the glucose PTS (deletion of ptsG, ptsH, ptsI, or crr) have been previously investigated (4,5,17,26,39,45). These mutants have a reduced growth rate in glucose minimal medium and exhibit a strongly reduced rate of acetate formation (45) even in complex media, where the growth rate is unaffected (4).…”

mentioning

confidence: 99%

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Glucose Transport in Escherichia coli Mutant Strains with Defects in Sugar Transport Systems

Steinsiek

Bettenbrock

2012

J Bacteriol

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In Escherichia coli, several systems are known to transport glucose into the cytoplasm. The main glucose uptake system under batch conditions is the glucose phosphoenolpyruvate:carbohydrate phosphotransferase system (glucose PTS), but the mannose PTS and the galactose and maltose transporters also can translocate glucose. Mutant strains which lack the enzyme IIBC (EIIBC) protein of the glucose PTS have been investigated previously because their lower rate of acetate formation offers advantages in industrial applications. Nevertheless, a systematic study to analyze the impact of the different glucose uptake systems has not been undertaken. Specifically, how the bacteria cope with the deletion of the major glucose uptake system and which alternative transporters react to compensate for this deficit have not been studied in detail. Therefore, a series of mutant strains were analyzed in aerobic and anaerobic batch cultures, as well as glucose-limited continuous cultivations. Deletion of EIIBC disturbs glucose transport severely in batch cultures; cyclic AMP (cAMP)-cAMP receptor protein (CRP) levels rise, and induction of the mgl operon occurs. Nevertheless, Mgl activity is not essential for growth of these mutants, since deletion of this transporter did not affect the growth rate; the activities of the remaining transporters seem to be sufficient. Under conditions of glucose limitation, mgl is upregulated 23-fold compared to levels for growth under glucose excess. Despite the strong induction of mgl upon glucose limitation, deletion of this transport system did not lead to further changes. Although the galactose transporters are often regarded as important for glucose uptake at micromolar concentrations, the glucose as well as mannose PTS might be sufficient for growth at this relatively low dilution rate.T he transport of carbon sources, especially glucose, is an important field of research that has been previously investigated in Escherichia coli. The main uptake system under conditions of glucose excess is the phosphoenolpyruvate:carbohydrate phosphotransferase system (PTS). It plays an important role not only in the transport and phosphorylation of carbon sources but also in the regulation of carbon metabolism and chemotaxis (for an overview, see reviews in references 11a, 47a, 50, and 50a). Glucose transport into the cytoplasm by EIICB Glc (encoded by ptsG) is coupled to its phosphorylation. The phosphate group is derived from phosphoenolpyruvate (PEP) and is transferred via a cascade of proteins, enzyme I (EI), HPr, EIIA, and EIIB. EI and HPr are general PTS proteins, while the EII proteins are carbohydrate specific. Like the glucose PTS, the mannose PTS (encoded by manXYZ) has been shown to play a role in transportation and phosphorylation of glucose under aerobic conditions with glucose as the sole carbon source (26), although PtsM has a lower affinity for glucose than PtsG (K m PtsM , 1.3 mM [9]; K m PtsG , 10 to 20 M [26]). Besides the phosphotransferase systems, the galactose transport systems, the galact...

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Section: Discussioncontrasting

confidence: 62%

Section: Discussionmentioning

confidence: 97%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Glucose Transport in Escherichia coli Mutant Strains with Defects in Sugar Transport Systems

Steinsiek

Bettenbrock

2012

J Bacteriol

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“…The resulting strains used have all mutations situated in the genes for expression of the phosphoenolpyruvate:carbohydrate phosphotransferase system (PTS) and results in a reduced uptake rate of glucose [4]. The three strains are defective in either the enzymes IIBC Glc (ptsG), IIAB Man (manX) or both.…”

Section: Strains and Vectorsmentioning

confidence: 99%

Cell engineering of Escherichia coli allows high cell density accumulation without fed-batch process control

Bäcklund

Markland

Larsson

2007

Bioprocess Biosyst Eng

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A set of mutations in the phosphoenolpyruvate:carbohydrate phosphotransferase system (PTS) was used to create Escherichia coli strains with a reduced uptake rate of glucose. This allows a growth restriction, which is controlled on cellular rather than reactor level, which is typical of the fed-batch cultivation concept. Batch growth of the engineered strains resulted in cell accumulation profiles corresponding to a growth rate of 0.78, 0.38 and 0.25 h(-1), respectively. The performance of the mutants in batch cultivation was compared to fed-batch cultivation of the wild type cell using restricted glucose feed to arrive at the corresponding growth profiles. Results show that the acetate production, oxygen consumption and product formation were similar, when a recombinant product was induced from the lacUV5 promoter. Ten times more cells could be produced in batch cultivation using the mutants without the growth detrimental production of acetic acid. This allows high cell density production without the establishment of elaborate fed-batch control equipment. The technique is suggested as a versatile tool in high throughput multiparallel protein production but also for increasing the number of experiments performed during process development while keeping conditions similar to the large-scale fed-batch performance.

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A hidden square‐root boundary between growth rate and biomass yield

Wong

Tran

Liao

2008

Biotech & Bioengineering

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Although the theoretical value of biomass yield can be calculated from metabolic network stoichiometry, the growth rate is difficult to predict. Since the rate and yield can vary independently, no simple relationship has been discovered between these two variables. In this work, we analyzed the well-accepted enzyme kinetics and uncovered a hidden boundary for growth rate, which is determined by the square-root of three physiological parameters: biomass yield, the substrate turnover number, and the maximum synthesis rate of the turnover enzyme. Cells cannot grow faster than the square-root of the product of these parameters. This analysis is supported by experimental data and involves essentially no assumptions except (i) the cell is not undergoing a downshift transition, (ii) substrate uptake enzyme activity is proportional to its copy number. This simple boundary (not correlation) has escaped notice for many decades and suggests that the yield calculation does not predict the growth rate, but gives an upper limit for the growth rate. The relationship also explains how growth rate is affected by the yield and sheds lights on strain design for product formation.

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Reducing the glucose uptake rate in Escherichia coli affects growth rate but not protein production

Cited by 40 publications

References 39 publications

Glucose Transport in Escherichia coli Mutant Strains with Defects in Sugar Transport Systems

Glucose Transport in Escherichia coli Mutant Strains with Defects in Sugar Transport Systems

Cell engineering of Escherichia coli allows high cell density accumulation without fed-batch process control

A hidden square‐root boundary between growth rate and biomass yield

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