Details
Originalsprache | Englisch |
---|---|
Seiten (von - bis) | 375-384 |
Seitenumfang | 10 |
Fachzeitschrift | Journal of biotechnology |
Jahrgang | 132 |
Ausgabenummer | 4 |
Publikationsstatus | Veröffentlicht - 1 Dez. 2007 |
Extern publiziert | Ja |
Abstract
The response of the central carbon metabolism of Escherichia coli to temperature-induced recombinant production of human fibroblast growth factor was studied on the level of metabolic fluxes and intracellular metabolite levels. During production, E. coli TG1:pλFGFB, carrying a plasmid encoded gene for the recombinant product, revealed stress related characteristics such as decreased growth rate and biomass yield and enhanced by-product excretion (acetate, pyruvate, lactate). With the onset of production, the adenylate energy charge dropped from 0.85 to 0.60, indicating the occurrence of a severe energy limitation. This triggered an increase of the glycolytic flux which, however, was not sufficient to compensate for the increased ATP demand. The activation of the glycolytic flux was also indicated by the readjustment of glycolytic pool sizes leading to an increased driving force for the reaction catalyzed by phosphofructokinase. Moreover, fluxes through the TCA cycle, into the pentose phosphate pathway and into anabolic pathways decreased significantly. The strong increase of flux into overflow pathways, especially towards acetate was most likely caused by a flux redirection from pyruvate dehydrogenase to pyruvate oxidase. The glyoxylate shunt, not active during growth, was the dominating anaplerotic pathway during production. Together with pyruvate oxidase and acetyl CoA synthase this pathway could function as a metabolic by-pass to overcome the limitation in the junction between glycolysis and TCA cycle and partly recycle the acetate formed back into the metabolism.
ASJC Scopus Sachgebiete
- Biochemie, Genetik und Molekularbiologie (insg.)
- Biotechnologie
- Chemische Verfahrenstechnik (insg.)
- Bioengineering
- Immunologie und Mikrobiologie (insg.)
- Angewandte Mikrobiologie und Biotechnologie
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in: Journal of biotechnology, Jahrgang 132, Nr. 4, 01.12.2007, S. 375-384.
Publikation: Beitrag in Fachzeitschrift › Artikel › Forschung › Peer-Review
}
TY - JOUR
T1 - Response of fluxome and metabolome to temperature-induced recombinant protein synthesis in Escherichia coli
AU - Wittmann, Christoph
AU - Weber, Jan
AU - Betiku, Eriola
AU - Krömer, Jens
AU - Böhm, Daniela
AU - Rinas, Ursula
N1 - Funding Information: Eriola Betiku gratefully acknowledges financial support by the Deutscher Akademischer Austauschdienst (DAAD). Jens Krömer acknowledges financial support of the BASF AG (Ludwigshafen, Germany).
PY - 2007/12/1
Y1 - 2007/12/1
N2 - The response of the central carbon metabolism of Escherichia coli to temperature-induced recombinant production of human fibroblast growth factor was studied on the level of metabolic fluxes and intracellular metabolite levels. During production, E. coli TG1:pλFGFB, carrying a plasmid encoded gene for the recombinant product, revealed stress related characteristics such as decreased growth rate and biomass yield and enhanced by-product excretion (acetate, pyruvate, lactate). With the onset of production, the adenylate energy charge dropped from 0.85 to 0.60, indicating the occurrence of a severe energy limitation. This triggered an increase of the glycolytic flux which, however, was not sufficient to compensate for the increased ATP demand. The activation of the glycolytic flux was also indicated by the readjustment of glycolytic pool sizes leading to an increased driving force for the reaction catalyzed by phosphofructokinase. Moreover, fluxes through the TCA cycle, into the pentose phosphate pathway and into anabolic pathways decreased significantly. The strong increase of flux into overflow pathways, especially towards acetate was most likely caused by a flux redirection from pyruvate dehydrogenase to pyruvate oxidase. The glyoxylate shunt, not active during growth, was the dominating anaplerotic pathway during production. Together with pyruvate oxidase and acetyl CoA synthase this pathway could function as a metabolic by-pass to overcome the limitation in the junction between glycolysis and TCA cycle and partly recycle the acetate formed back into the metabolism.
AB - The response of the central carbon metabolism of Escherichia coli to temperature-induced recombinant production of human fibroblast growth factor was studied on the level of metabolic fluxes and intracellular metabolite levels. During production, E. coli TG1:pλFGFB, carrying a plasmid encoded gene for the recombinant product, revealed stress related characteristics such as decreased growth rate and biomass yield and enhanced by-product excretion (acetate, pyruvate, lactate). With the onset of production, the adenylate energy charge dropped from 0.85 to 0.60, indicating the occurrence of a severe energy limitation. This triggered an increase of the glycolytic flux which, however, was not sufficient to compensate for the increased ATP demand. The activation of the glycolytic flux was also indicated by the readjustment of glycolytic pool sizes leading to an increased driving force for the reaction catalyzed by phosphofructokinase. Moreover, fluxes through the TCA cycle, into the pentose phosphate pathway and into anabolic pathways decreased significantly. The strong increase of flux into overflow pathways, especially towards acetate was most likely caused by a flux redirection from pyruvate dehydrogenase to pyruvate oxidase. The glyoxylate shunt, not active during growth, was the dominating anaplerotic pathway during production. Together with pyruvate oxidase and acetyl CoA synthase this pathway could function as a metabolic by-pass to overcome the limitation in the junction between glycolysis and TCA cycle and partly recycle the acetate formed back into the metabolism.
KW - C metabolic flux
KW - Adenylate energy charge
KW - Cellular stress
KW - Human basic fibroblast growth factor
UR - http://www.scopus.com/inward/record.url?scp=35748959651&partnerID=8YFLogxK
U2 - 10.1016/j.jbiotec.2007.07.495
DO - 10.1016/j.jbiotec.2007.07.495
M3 - Article
C2 - 17689798
AN - SCOPUS:35748959651
VL - 132
SP - 375
EP - 384
JO - Journal of biotechnology
JF - Journal of biotechnology
SN - 0168-1656
IS - 4
ER -