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Physiological and Transcriptional Characterization of Escherichia Coli Strains Lacking Interconversion of Phosphoenolpyruvate and Pyruvate When Glucose and Acetate are Coutilized

Phosphoenolpyruvate (PEP) is a precursor involved in the biosynthesis of aromatics and other valuable compounds in Escherichia coli. The PEP:carbohydrate phosphotransferase system (PTS) is the major glucose transport system and the largest PEP consumer. To increase intracellular PEP availability for...

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Autores principales: Sabido, Andrea, Sigala, Juan Carlos, Hernández-Chávez, Georgina, Flores, Noemí, Gosset, Guillermo, Bolívar, Francisco
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BlackWell Publishing Ltd 2013
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4278548/
https://www.ncbi.nlm.nih.gov/pubmed/24375081
http://dx.doi.org/10.1002/bit.25177
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author Sabido, Andrea
Sigala, Juan Carlos
Hernández-Chávez, Georgina
Flores, Noemí
Gosset, Guillermo
Bolívar, Francisco
author_facet Sabido, Andrea
Sigala, Juan Carlos
Hernández-Chávez, Georgina
Flores, Noemí
Gosset, Guillermo
Bolívar, Francisco
author_sort Sabido, Andrea
collection PubMed
description Phosphoenolpyruvate (PEP) is a precursor involved in the biosynthesis of aromatics and other valuable compounds in Escherichia coli. The PEP:carbohydrate phosphotransferase system (PTS) is the major glucose transport system and the largest PEP consumer. To increase intracellular PEP availability for aromatics production purposes, mutant strains of E. coli JM101 devoid of the ptsHIcrr operon (PB11 strain) have been previously generated. In this derivative, transport and growth rate on glucose decreased significantly. A laboratory evolved strain derived from PB11 that partially recovered its growth capacity on glucose was named PB12. In the present study, we blocked carbon skeletons interchange between PEP and pyruvate (PYR) in these ptsHIcrr(−) strains by deleting the pykA, pykF, and ppsA genes. The PB11 pykAF(−) ppsA(−) strain exhibited no growth on glucose or acetate alone, but it was viable when both substrates were consumed simultaneously. In contrast, the PB12 pykAF(−) ppsA(−) strain displayed a low growth rate on glucose or acetate alone, but in the mixture, growth was significantly improved. RT-qPCR expression analysis of PB11 pykAF(−) ppsA(−) growing with both carbon sources showed a downregulation of all central metabolic pathways compared with its parental PB11 strain. Under the same conditions, transcription of most of the genes in PB12 pykAF(−) ppsA(−) did not change, and few like aceBAK, sfcA, and poxB were overexpressed compared with PB12. We explored the aromatics production capabilities of both ptsHIcrr(−) pykAF(−) ppsA(−) strains and the engineered PB12 pykAF(−) ppsA(−) tyrR(−) pheA(ev2+)/pJLBaroG(fbr)tktA enhanced the yield of aromatic compounds when coutilizing glucose and acetate compared with the control strain PB12 tyrR(−) pheA(ev2+)/pJLBaroG(fbr)tktA. Biotechnol. Bioeng. 2014;111: 1150–1160. © 2013 The Authors. Biotechnology and Bioengineering Published by Wiley Periodicals, Inc.
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spelling pubmed-42785482014-12-30 Physiological and Transcriptional Characterization of Escherichia Coli Strains Lacking Interconversion of Phosphoenolpyruvate and Pyruvate When Glucose and Acetate are Coutilized Sabido, Andrea Sigala, Juan Carlos Hernández-Chávez, Georgina Flores, Noemí Gosset, Guillermo Bolívar, Francisco Biotechnol Bioeng Articles Phosphoenolpyruvate (PEP) is a precursor involved in the biosynthesis of aromatics and other valuable compounds in Escherichia coli. The PEP:carbohydrate phosphotransferase system (PTS) is the major glucose transport system and the largest PEP consumer. To increase intracellular PEP availability for aromatics production purposes, mutant strains of E. coli JM101 devoid of the ptsHIcrr operon (PB11 strain) have been previously generated. In this derivative, transport and growth rate on glucose decreased significantly. A laboratory evolved strain derived from PB11 that partially recovered its growth capacity on glucose was named PB12. In the present study, we blocked carbon skeletons interchange between PEP and pyruvate (PYR) in these ptsHIcrr(−) strains by deleting the pykA, pykF, and ppsA genes. The PB11 pykAF(−) ppsA(−) strain exhibited no growth on glucose or acetate alone, but it was viable when both substrates were consumed simultaneously. In contrast, the PB12 pykAF(−) ppsA(−) strain displayed a low growth rate on glucose or acetate alone, but in the mixture, growth was significantly improved. RT-qPCR expression analysis of PB11 pykAF(−) ppsA(−) growing with both carbon sources showed a downregulation of all central metabolic pathways compared with its parental PB11 strain. Under the same conditions, transcription of most of the genes in PB12 pykAF(−) ppsA(−) did not change, and few like aceBAK, sfcA, and poxB were overexpressed compared with PB12. We explored the aromatics production capabilities of both ptsHIcrr(−) pykAF(−) ppsA(−) strains and the engineered PB12 pykAF(−) ppsA(−) tyrR(−) pheA(ev2+)/pJLBaroG(fbr)tktA enhanced the yield of aromatic compounds when coutilizing glucose and acetate compared with the control strain PB12 tyrR(−) pheA(ev2+)/pJLBaroG(fbr)tktA. Biotechnol. Bioeng. 2014;111: 1150–1160. © 2013 The Authors. Biotechnology and Bioengineering Published by Wiley Periodicals, Inc. BlackWell Publishing Ltd 2013-06 2013-01-28 /pmc/articles/PMC4278548/ /pubmed/24375081 http://dx.doi.org/10.1002/bit.25177 Text en © 2013 The Authors. Biotechnology and Bioengineering Published by Wiley Periodicals, Inc. http://creativecommons.org/licenses/by-nc-nd/3.0/ This is an open access article under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made.
spellingShingle Articles
Sabido, Andrea
Sigala, Juan Carlos
Hernández-Chávez, Georgina
Flores, Noemí
Gosset, Guillermo
Bolívar, Francisco
Physiological and Transcriptional Characterization of Escherichia Coli Strains Lacking Interconversion of Phosphoenolpyruvate and Pyruvate When Glucose and Acetate are Coutilized
title Physiological and Transcriptional Characterization of Escherichia Coli Strains Lacking Interconversion of Phosphoenolpyruvate and Pyruvate When Glucose and Acetate are Coutilized
title_full Physiological and Transcriptional Characterization of Escherichia Coli Strains Lacking Interconversion of Phosphoenolpyruvate and Pyruvate When Glucose and Acetate are Coutilized
title_fullStr Physiological and Transcriptional Characterization of Escherichia Coli Strains Lacking Interconversion of Phosphoenolpyruvate and Pyruvate When Glucose and Acetate are Coutilized
title_full_unstemmed Physiological and Transcriptional Characterization of Escherichia Coli Strains Lacking Interconversion of Phosphoenolpyruvate and Pyruvate When Glucose and Acetate are Coutilized
title_short Physiological and Transcriptional Characterization of Escherichia Coli Strains Lacking Interconversion of Phosphoenolpyruvate and Pyruvate When Glucose and Acetate are Coutilized
title_sort physiological and transcriptional characterization of escherichia coli strains lacking interconversion of phosphoenolpyruvate and pyruvate when glucose and acetate are coutilized
topic Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4278548/
https://www.ncbi.nlm.nih.gov/pubmed/24375081
http://dx.doi.org/10.1002/bit.25177
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