Stationary versus non-stationary 13C-MFA: A comparison using a consistent dataset

Noack, Stephan; Nöh, Katharina; Moch, Matthias; Oldiges, Marco; Wiechert, Wolfgang

doi:10.1016/j.jbiotec.2010.07.008

Cited by 57 publications

(42 citation statements)

References 55 publications

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“…As already shown in former experiments following the labeling of transients in a C. glutamicum L-lysine producer and E. coli wild type (25,26), isotopic stationarity is rapidly achieved (i.e., within a few minutes) for intermediates of the EMP and PPP. In contrast, the time constants for TCA cycle intermediates are much higher, leading to significantly delays in labeling dynamics; isotopic stationary states are not reached until 1 h of labeling.…”

Section: Discussionsupporting

confidence: 60%

“…The calculation of intracellular reaction rates presented here is based on the classical 13 C MFA approach that strictly relies on the assumption of isotopic stationarity (25). As already shown in former experiments following the labeling of transients in a C. glutamicum L-lysine producer and E. coli wild type (25,26), isotopic stationarity is rapidly achieved (i.e., within a few minutes) for intermediates of the EMP and PPP.…”

Section: Discussionmentioning

confidence: 99%

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Comparative¹³C Metabolic Flux Analysis of Pyruvate Dehydrogenase Complex-Deficient,l-Valine-Producing Corynebacterium glutamicum

Bartek

Blombach

Lang

et al. 2011

Appl Environ Microbiol

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L-Valine can be formed successfully using C. glutamicum strains missing an active pyruvate dehydrogenase enzyme complex (PDHC). Wild-type C. glutamicum and four PDHC-deficient strains were compared by 13 C metabolic flux analysis, especially focusing on the split ratio between glycolysis and the pentose phosphate pathway (PPP). Compared to the wild type, showing a carbon flux of 69% ؎ 14% through the PPP, a strong increase in the PPP flux was observed in PDHC-deficient strains with a maximum of 113% ؎ 22%. The shift in the split ratio can be explained by an increased demand of NADPH for L-valine formation. In accordance, the introduction of the Escherichia coli transhydrogenase PntAB, catalyzing the reversible conversion of NADH to NADPH, into an L-valine-producing C. glutamicum strain caused the PPP flux to decrease to 57% ؎ 6%, which is below the wild-type split ratio. Hence, transhydrogenase activity offers an alternative perspective for sufficient NADPH supply, which is relevant for most amino acid production systems. Moreover, as demonstrated for L-valine, this bypass leads to a significant increase of product yield due to a concurrent reduction in carbon dioxide formation via the PPP.

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Section: Discussionsupporting

confidence: 60%

Section: Discussionmentioning

confidence: 99%

Comparative¹³C Metabolic Flux Analysis of Pyruvate Dehydrogenase Complex-Deficient,l-Valine-Producing Corynebacterium glutamicum

Bartek

Blombach

Lang

et al. 2011

Appl Environ Microbiol

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“…Standardization is also a prerequisite for comparability and reproducibility of fluxomics studies, which is a neglected aspect ( [66,75] are rare exceptions). The fundamental question is: how accurate and precise is the flux measurement?…”

Section: Discussionmentioning

confidence: 99%

How to measure metabolic fluxes: a taxonomic guide for 13 C fluxomics

Niedenführ

Wiechert

Nöh

2015

Current Opinion in Biotechnology

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113

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“…Therefore the additional measurements of labeling over time in a temporal analysis (Fig. 3) provide a richer set of data for modeling purposes [321] over steady state analysis alone. Additionally, because the earliest time points are the most sensitive to label provision, they provide a significant amount of information and the experimental duration can be shortened [322,323].…”

Section: From Co 2 To Lipid: Temporal Labeling-based Mfa Approachesmentioning

confidence: 99%

Tracking the metabolic pulse of plant lipid production with isotopic labeling and flux analyses: Past, present and future

Allen

Bates

Tjellström

2015

Progress in Lipid Research

100

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Metabolism is comprised of networks of chemical transformations, organized into integrated biochemical pathways that are the basis of cellular operation, and function to sustain life. Metabolism, and thus life, is not static. The rate of metabolites transitioning through biochemical pathways (i.e., flux) determines cellular phenotypes, and is constantly changing in response to genetic or environmental perturbations. Each change evokes a response in metabolic pathway flow, and the quantification of fluxes under varied conditions helps to elucidate major and minor routes, and regulatory aspects of metabolism. To measure fluxes requires experimental methods that assess the movements and transformations of metabolites without creating artifacts. Isotopic labeling fills this role and is a long-standing experimental approach to identify pathways and quantify their metabolic relevance in different tissues or under different conditions. The application of labeling techniques to plant science is however far from reaching it potential. In light of advances in genetics and molecular biology that provide a means to alter metabolism, and given recent improvements in instrumentation, computational tools and available isotopes, the use of isotopic labeling to probe metabolism is becoming more and more powerful. We review the principal analytical methods for isotopic labeling with a focus on seminal studies of pathways and fluxes in lipid metabolism and carbon partitioning through central metabolism. Central carbon metabolic steps are directly linked to lipid production by serving to generate the precursors for fatty acid biosynthesis and lipid assembly. Additionally some of the ideas for labeling techniques that may be most applicable for lipid metabolism in the future were originally developed to investigate other aspects of central metabolism. We conclude by describing recent advances that will play an important future role in quantifying flux and metabolic operation in plant tissues.

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Stationary versus non-stationary 13C-MFA: A comparison using a consistent dataset

Cited by 57 publications

References 55 publications

Comparative¹³C Metabolic Flux Analysis of Pyruvate Dehydrogenase Complex-Deficient,l-Valine-Producing Corynebacterium glutamicum

Comparative¹³C Metabolic Flux Analysis of Pyruvate Dehydrogenase Complex-Deficient,l-Valine-Producing Corynebacterium glutamicum

How to measure metabolic fluxes: a taxonomic guide for 13 C fluxomics

Tracking the metabolic pulse of plant lipid production with isotopic labeling and flux analyses: Past, present and future

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Stationary versus non-stationary 13C-MFA: A comparison using a consistent dataset

Cited by 57 publications

References 55 publications

Comparative13C Metabolic Flux Analysis of Pyruvate Dehydrogenase Complex-Deficient,l-Valine-Producing Corynebacterium glutamicum

Comparative13C Metabolic Flux Analysis of Pyruvate Dehydrogenase Complex-Deficient,l-Valine-Producing Corynebacterium glutamicum

How to measure metabolic fluxes: a taxonomic guide for 13 C fluxomics

Tracking the metabolic pulse of plant lipid production with isotopic labeling and flux analyses: Past, present and future

Contact Info

Product

Resources

About

Comparative¹³C Metabolic Flux Analysis of Pyruvate Dehydrogenase Complex-Deficient,l-Valine-Producing Corynebacterium glutamicum

Comparative¹³C Metabolic Flux Analysis of Pyruvate Dehydrogenase Complex-Deficient,l-Valine-Producing Corynebacterium glutamicum