Pseudomonas aeruginosa is an important platform for simultaneous
production of polyhydroxyalkanoates (PHA) and rhamnolipids (RHL).
Advances in the understanding of the biosynthesis metabolism of these
biocompounds are crucial for increasing yield. 13C-Metabolic Flux Ratio
Analysis (13C-MFA) is a technique to estimate in vivo metabolic fluxes
ratios. PHA and RHL are essentially non-growth associated products of
biotechnological interest and both contain hydroxyalkanoates (HAs),
whose labeling patterns could be accessed by GC-MS. In this study, to
reveal the relative contributions of the Entner-Doudoroff (ED) pathway
and the non-oxidative Pentose Phosphate (PP) pathway to PHA and RHL
production, 13C-MFA was performed in Pseudomonas aeruginosa LFM634 when
supplied with labeled glucose. This strain lacks both functional EMP and
the oxidative PP branch. Labeling patterns in HAs were measured.
Experiments with [U-13C] glucose indicated a low flux trough PP
pathway. An optimal design of labeling experiment showed that
[6-13C] glucose would be the best substrate to enable an estimation
of the ED flux with high accuracy. Results of experiments performed with
this isotope indicated that about two-thirds of glyceraldehyde
3-phosphate is recycled through a cyclic ED architecture, suggesting
that P. aeruginosa utilizes that cycle to regulate the NADPH/Acetyl-CoA
ratio for PHA and RHL biosynthesis.