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Modification of glucose import capacity in Escherichia coli: physiologic consequences and utility for improving DNA vaccine production.
Microb Cell Fact. 2013 May 02; 12:42.MC

Abstract

BACKGROUND

The bacterium Escherichia coli can be grown employing various carbohydrates as sole carbon and energy source. Among them, glucose affords the highest growth rate. This sugar is nowadays widely employed as raw material in industrial fermentations. When E. coli grows in a medium containing non-limiting concentrations of glucose, a metabolic imbalance occurs whose main consequence is acetate secretion. The production of this toxic organic acid reduces strain productivity and viability. Solutions to this problem include reducing glucose concentration by substrate feeding strategies or the generation of mutant strains with impaired glucose import capacity. In this work, a collection of E. coli strains with inactive genes encoding proteins involved in glucose transport where generated to determine the effects of reduced glucose import capacity on growth rate, biomass yield, acetate and production of an experimental plasmid DNA vaccine (pHN).

RESULTS

A group of 15 isogenic derivatives of E. coli W3110 were generated with single and multiple deletions of genes encoding glucose, mannose, beta-glucoside, maltose and N-acetylglucosamine components of the phosphoenolpyruvate:sugar phosphotransferase system (PTS), as well as the galactose symporter and the Mgl galactose/glucose ABC transporter. These strains were characterized by growing them in mineral salts medium supplemented with 2.5 g/L glucose. Maximum specific rates of glucose consumption (qs) spanning from 1.33 to 0.32 g/g h were displayed by the group of mutants and W3110, which resulted in specific growth rates ranging from 0.65-0.18 h(-1). Acetate accumulation was reduced or abolished in cultures with all mutant strains. W3110 and five selected mutant derivatives were transformed with pHN. A 3.2-fold increase in pHN yield on biomass was observed in cultures of a mutant strain with deletion of genes encoding the glucose and mannose PTS components, as well as Mgl.

CONCLUSIONS

The group of E. coli mutants generated in this study displayed a reduction or elimination of overflow metabolism and a linear correlation between qs and the maximum specific growth rate as well as the acetate production rate. By comparing DNA vaccine production parameters among some of these mutants, it was possible to identify a near-optimal glucose import rate value for this particular application. The strains employed in this study should be a useful resource for studying the effects of different predefined qs values on production capacity for various biotechnological products.

Authors+Show Affiliations

Departamento de Ingeniería Celular y Biocatálisis, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Cuernavaca, Morelos, Mexico.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

23638701

Citation

Fuentes, Laura G., et al. "Modification of Glucose Import Capacity in Escherichia Coli: Physiologic Consequences and Utility for Improving DNA Vaccine Production." Microbial Cell Factories, vol. 12, 2013, p. 42.
Fuentes LG, Lara AR, Martínez LM, et al. Modification of glucose import capacity in Escherichia coli: physiologic consequences and utility for improving DNA vaccine production. Microb Cell Fact. 2013;12:42.
Fuentes, L. G., Lara, A. R., Martínez, L. M., Ramírez, O. T., Martínez, A., Bolívar, F., & Gosset, G. (2013). Modification of glucose import capacity in Escherichia coli: physiologic consequences and utility for improving DNA vaccine production. Microbial Cell Factories, 12, 42. https://doi.org/10.1186/1475-2859-12-42
Fuentes LG, et al. Modification of Glucose Import Capacity in Escherichia Coli: Physiologic Consequences and Utility for Improving DNA Vaccine Production. Microb Cell Fact. 2013 May 2;12:42. PubMed PMID: 23638701.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Modification of glucose import capacity in Escherichia coli: physiologic consequences and utility for improving DNA vaccine production. AU - Fuentes,Laura G, AU - Lara,Alvaro R, AU - Martínez,Luz M, AU - Ramírez,Octavio T, AU - Martínez,Alfredo, AU - Bolívar,Francisco, AU - Gosset,Guillermo, Y1 - 2013/05/02/ PY - 2013/03/13/received PY - 2013/04/26/accepted PY - 2013/5/4/entrez PY - 2013/5/4/pubmed PY - 2013/12/16/medline SP - 42 EP - 42 JF - Microbial cell factories JO - Microb. Cell Fact. VL - 12 N2 - BACKGROUND: The bacterium Escherichia coli can be grown employing various carbohydrates as sole carbon and energy source. Among them, glucose affords the highest growth rate. This sugar is nowadays widely employed as raw material in industrial fermentations. When E. coli grows in a medium containing non-limiting concentrations of glucose, a metabolic imbalance occurs whose main consequence is acetate secretion. The production of this toxic organic acid reduces strain productivity and viability. Solutions to this problem include reducing glucose concentration by substrate feeding strategies or the generation of mutant strains with impaired glucose import capacity. In this work, a collection of E. coli strains with inactive genes encoding proteins involved in glucose transport where generated to determine the effects of reduced glucose import capacity on growth rate, biomass yield, acetate and production of an experimental plasmid DNA vaccine (pHN). RESULTS: A group of 15 isogenic derivatives of E. coli W3110 were generated with single and multiple deletions of genes encoding glucose, mannose, beta-glucoside, maltose and N-acetylglucosamine components of the phosphoenolpyruvate:sugar phosphotransferase system (PTS), as well as the galactose symporter and the Mgl galactose/glucose ABC transporter. These strains were characterized by growing them in mineral salts medium supplemented with 2.5 g/L glucose. Maximum specific rates of glucose consumption (qs) spanning from 1.33 to 0.32 g/g h were displayed by the group of mutants and W3110, which resulted in specific growth rates ranging from 0.65-0.18 h(-1). Acetate accumulation was reduced or abolished in cultures with all mutant strains. W3110 and five selected mutant derivatives were transformed with pHN. A 3.2-fold increase in pHN yield on biomass was observed in cultures of a mutant strain with deletion of genes encoding the glucose and mannose PTS components, as well as Mgl. CONCLUSIONS: The group of E. coli mutants generated in this study displayed a reduction or elimination of overflow metabolism and a linear correlation between qs and the maximum specific growth rate as well as the acetate production rate. By comparing DNA vaccine production parameters among some of these mutants, it was possible to identify a near-optimal glucose import rate value for this particular application. The strains employed in this study should be a useful resource for studying the effects of different predefined qs values on production capacity for various biotechnological products. SN - 1475-2859 UR - https://www.unboundmedicine.com/medline/citation/23638701/Modification_of_glucose_import_capacity_in_Escherichia_coli:_physiologic_consequences_and_utility_for_improving_DNA_vaccine_production_ L2 - https://microbialcellfactories.biomedcentral.com/articles/10.1186/1475-2859-12-42 DB - PRIME DP - Unbound Medicine ER -