FiatFlux-P - User-Level Workflow Design: A Bioinformatics Perspective - page 100

Fig. 5.1 Metabolic flux analysis based on 13 C labeling experiments
methods exists that can identify and quantify defined sets of compounds in
parallel. The most frequently applied methods for metabolic profiling are nu-
clear magnetic resonance (NMR) and mass spectrometry (MS), where the lat-
ter is usually preceded by a chromatographic step (such as gas chromatogra-
phy ,GC,or liquid chromatography , LC) to separate the metabolites. Knowledge
from metabolomics experiments can be used for building or refining models of
metabolic pathways. As metabolites are at the end of the cellular information
chain [279, p. 165 ff.], their quantification also allows to draw inferences about
other cell networks, such as gene regulatory networks or signal transduction
pathways.
Figure 5.1 sketches the procedure of a so-called 13 C tracer experiment :
Glucose, a sugar consisting of six carbon atoms, is labeled with a 13 Cisotope
at the first carbon atom of the sugar molecule (upper left of the figure). The
marked glucose is then fed to the biological system and, due to the catabolism
of the sugar molecule, the labeled atoms are distributed over the metabolic
network. Later on, the labeling patterns of intracellular metabolites or amino
acids located in proteins can be measured by NMR or MS instruments. The
resulting spectra provide information about the physiology of the biological
system under investigation. Depending on the organism, there are several pos-
sibilities of how the cell processes the glucose into pyruvate (three C atoms),
with the purpose of gaining energy from the molecule splitting. The microor-
ganism Escherichia coli, for instance can metabolize glucose via the Entner-
Doudoroff pathway ,the glycolysis ,andthe Pentose-Phosphate pathway .As
the figure (upper left) shows, the Entner-Doudoroff pathway results in two
pyruvate molecules of which is unlabeled and the second contains the 13 Cof
the glucose at the first carbon position. Glucose catabolism via the glycolysis
results in a 50:50 mixture of unlabeled pyruvate and molecules carrying the
13 C isotope at the C 3 position. When glucose is processed via the Pentose-
phosphate pathway, neither of the resulting pyruvate molecules contains a
13 C atom. In the experiment, the labeled glucose is fed to an organism, and
the resulting pyruvate molecules are measured and classified according to the