Journal article
Combined Rapid Injection NMR and Simulation Approach to Probe Redox-Dependent Pathway Control in Living Cells
Magnetic Resonance, Department of Health Technology, Technical University of Denmark1
Department of Health Technology, Technical University of Denmark2
Center for Hyperpolarization in Magnetic Resonance, Centers, Technical University of Denmark3
Quantitative Modeling of Cell Metabolism, Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark4
iLoop, Translational Management, Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark5
Research Groups, Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark6
Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark7
Department of Chemistry, Technical University of Denmark8
Dynamic response of intracellular reaction cascades to changing environments is a hallmark of living systems. As metabolism is complex, mechanistic models have gained popularity for describing the dynamic response of cellular metabolism and for identifying target genes for engineering. At the same time, the detailed tracking of transient metabolism in living cells on the sub-minute timescale has become amenable using dynamic nuclear polarization enhanced 13C NMR.
Here, we suggest an approach combining in-cell NMR spectroscopy with perturbation experiments and modeling to obtain evidence that the bottlenecks of yeast glycolysis depend on intracellular redox state. In pre-steady state glycolysis, pathway bottlenecks shift from downstream to upstream reactions within few seconds, consistent with a rapid decline in the NAD+/NADH ratio.
Simulations using mechanistic models reproduce the experimentally observed response and help identify unforeseen biochemical events. Remaining inaccuracies in the computational models can be identified experimentally. The combined use of rapid injection NMR spectroscopy and in silico simulations provides a promising method for characterizing cellular reactions with increasing mechanistic detail.
Language: | English |
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Publisher: | American Chemical Society |
Year: | 2019 |
Pages: | 5395-5402 |
ISSN: | 15204782 , 00032700 and 15206882 |
Types: | Journal article |
DOI: | 10.1021/acs.analchem.9b00660 |
ORCIDs: | Jensen, Pernille Rose , Matos, Marta , Sonnenschein, Nikolaus and Meier, Sebastian |