The availability of accurate reaction atom mapping models describing carbon flows between central and peripheral metabolism remains a major bottleneck in the elucidation of fluxes in photosynthetic cyanobacteria. In addition to altering predicted 13C-labeling distributions as shown in E. coli, a comprehensive carbon mapping model representing photoautotrophic growth affects labeling dynamics of all intracellular metabolites, which is vital to accurate flux resolution using isotopic non-stationary 13C-flux analysis. Here we highlight the construction of genome-scale metabolic mapping models imSyn711 and imSyf608 describing reaction atom mapping for all carbon-balanced reactions in cyanobacteria Synechocystis PCC 6803 and Synechococcus PCC 7942. Synechococcus and Synechocystis share 591 reactions with 120 reactions unique to Synechocystis and 17 unique to Synechococcus. Inherent differences in metabolism surrounding 3-phosphoglycerate (3PG), and glyceraldehyde-3-phosphate (G3P) are revealed to be the primary source of various carbon scrambling patterns unique to E. coli and cyanobacteria. The interplay between enzymes of the non-oxidative pentose phosphate pathway and carbon fixation via the Calvin cycle results in a rearrangement of C1 and C2 of the pentose phosphates, facilitating the transfer of fixed 13C-CO2 to all atoms of G3P and 3PG. In contrast, scrambling of such carbons occurs via upper glycolysis, pentose phosphate pathway, and the ED pathway, thereby limiting the extent of 13C-label distribution in E. coli. Furthermore, cyanobacterial mapping models reveal the presence of an additional 13C incorporation path via glyoxylate metabolism and photorespiration, absent in E. coli. Analysis of peripheral metabolism revealed that acetate can be a byproduct of porphyrin biosynthesis in cyanobacteria thus carrying additional labeling information upon reentry into central metabolism. iSyn711 is validated using genome-scale isotopic non-stationary MFA, which quantifies 681 intracellular fluxes and 203 pool sizes. In addition to better recapitulation of experimentally observed labeling distributions of all measured central metabolites, the model predicts the existence of a serine biosynthesis route from 3PG and trace flux through the GABA shunt. The implications of these findings in the context of cyanobacterial biology and metabolic engineering are discussed.

Cyanobacterial genome-scale carbon mapping models for genome-scale 13C-metabolic flux elucidation

253rd National Meeting (2017)

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