In vivo investigations of glucose transport in Saccharomyces cerevisiae

Rizzi, Manfred; Theobald, Uwe; Querfurth, Erich; Rohrhirsch, Thilo; Baltes, Michael; Reuß, Matthias

doi:10.1002/(sici)1097-0290(19960205)49:3<316::aid-bit10>3.0.co;2-c

Cited by 56 publications

(29 citation statements)

References 39 publications

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“…Candidate metabolites that inhibit the facilitated diffusion of glucose are glucose (36) and G6P (2,31), with the latter showing a strong increase after the pulse. We cannot exclude, however, the inhibitory effect of intracellular glucose, as it was not measured.…”

Section: Discussionmentioning

confidence: 99%

Short-Term Metabolome Dynamics and Carbon, Electron, and ATP Balances in Chemostat-Grown Saccharomyces cerevisiae CEN.PK 113-7D following a Glucose Pulse

Dam

Schipper

et al. 2006

Appl Environ Microbiol

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The in vivo kinetics in Saccharomyces cerevisiae CEN.PK 113-7D was evaluated during a 300-second transient period after applying a glucose pulse to an aerobic, carbon-limited chemostat culture. We quantified the responses of extracellular metabolites, intracellular intermediates in primary metabolism, intracellular free amino acids, and in vivo rates of O 2 uptake and CO 2 evolution. With these measurements, dynamic carbon, electron, and ATP balances were set up to identify major carbon, electron, and energy sinks during the postpulse period. There were three distinct metabolic phases during this time. In phase I (0 to 50 seconds after the pulse), the carbon/electron balances closed up to 85%. The accumulation of glycolytic and storage compounds accounted for 60% of the consumed glucose, caused an energy depletion, and may have led to a temporary decrease in the anabolic flux. In phase II (50 to 150 seconds), the fermentative metabolism gradually became the most important carbon/electron sink. In phase III (150 to 300 seconds), 29% of the carbon uptake was not identified in the measurements, and the ATP balance had a large surplus. These results indicate an increase in the anabolic flux, which is consistent with macroscopic balances of extracellular fluxes and the observed increase in CO 2 evolution associated with nonfermentative metabolism. The identified metabolic processes involving major carbon, electron, and energy sinks must be taken into account in in vivo kinetic models based on short-term dynamic metabolome responses.Mathematical models of in vivo enzyme kinetics in microorganisms are important for understanding metabolic control mechanisms operating on the level of the metabolome and can be used to assist the rational redesign of metabolic pathways to enhance desired functionalities of microbes (54). Kinetic parameters in this kind of model can be obtained by stimulusresponse experiments, in which cells grown in a (quasi-)steady state are perturbed by an external stimulus and the dynamic responses of intra-and extracellular metabolites are monitored. The time window of observation is usually within tens to a few hundred seconds after the application of the stimulus, and the responses are usually attributed to rapid (allosteric) enzyme-metabolite interactions (38). Kinetic parameters can be estimated from the measured responses, based on a set of (dynamic) material balances (7, 30, 49), as follows: dx/dt ϭ Sv (equation 1), where x is a vector of the metabolite concentrations, S is the stoichiometry matrix, and v is a vector of the reaction rates as a function of (yet) unknown kinetic parameters. The stimulus-response methodology is an ideal tool for obtaining kinetic information and has been applied to various microorganisms under different growth conditions (7,26,32,38,52).For Saccharomyces cerevisiae, a frequently applied perturbation is the addition of a concentrated glucose solution, i.e., a glucose pulse, to a glucose-limited chemostat culture, thereby inducing a short-term Crabtree effect. Theoba...

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Section: Discussionmentioning

confidence: 99%

Short-Term Metabolome Dynamics and Carbon, Electron, and ATP Balances in Chemostat-Grown Saccharomyces cerevisiae CEN.PK 113-7D following a Glucose Pulse

Dam

Schipper

et al. 2006

Appl Environ Microbiol

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show abstract

“…Glucose transport into the cell occurs via a permease at the cell membrane, which is inhibited by g6p [67]. We have adopted the rate equation of [14].…”

Section: Glucose Transporter (Glut R2)mentioning

confidence: 99%

“…As a first step to assess the validity of the model, we tried to fit another set of steady-state of Rizzi et al [67] and glucose pulse data of Theobald et al [83] measured under different conditions, solely by changing expression levels of enzymes (Vmax), but no other parameters. The experimental conditions differed in the yeast strain used, the dilution rate, extracellular glucose concentration and ethanol supply in the medium.…”

Section: Fitting Additional Glucose Pulse Experimentsmentioning

confidence: 99%

A new model for the aerobic metabolism of yeast allows the detailed analysis of the metabolic regulation during glucose pulse

Kesten

Kummer

Sahle

et al. 2015

Biophysical Chemistry

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“…The culture broth could be sampled less than every second, followed by quenching and extraction procedures. Initially this technique was applied to yeast cultivation [358,359], and later it was applied to E. coli [360]. More recently, in vivo kinetics with rapid perturbation experiments have been investigated using the so-called BioScope [361].…”

Section: The Systems Biology Approachmentioning

confidence: 99%

Toward systematic metabolic engineering based on the analysis of metabolic regulation by the integration of different levels of information

Shimizu

2009

Biochemical Engineering Journal

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In vivo investigations of glucose transport in Saccharomyces cerevisiae

Cited by 56 publications

References 39 publications

Short-Term Metabolome Dynamics and Carbon, Electron, and ATP Balances in Chemostat-Grown Saccharomyces cerevisiae CEN.PK 113-7D following a Glucose Pulse

Short-Term Metabolome Dynamics and Carbon, Electron, and ATP Balances in Chemostat-Grown Saccharomyces cerevisiae CEN.PK 113-7D following a Glucose Pulse

A new model for the aerobic metabolism of yeast allows the detailed analysis of the metabolic regulation during glucose pulse

Toward systematic metabolic engineering based on the analysis of metabolic regulation by the integration of different levels of information

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