Metabolic flux analysis of heterotrophic growth in Chlamydomonas reinhardtii

Despite the wealth of knowledge available for C. reinhardtii, the central metabolic fluxes of growth on acetate have not yet been determined. In this study, (13)C-metabolic flux analysis ((13)C-MFA) was used to determine and quantify the metabolic pathways of primary metabolism in C. reinhardtii cells grown under heterotrophic conditions with acetate as the sole carbon source. Isotopic labeling patterns of compartment specific biomass derived metabolites were used to calculate the fluxes. It was found that acetate is ligated with coenzyme A in the three subcellular compartments (cytosol, mitochondria and plastid) included in the model. Two citrate synthases were found to potentially be involved in acetyl-coA metabolism; one localized in the mitochondria and the other acting outside the mitochondria. Labeling patterns demonstrate that Acetyl-coA synthesized in the plastid is directly incorporated in synthesis of fatty acids. Despite having a complete TCA cycle in the mitochondria, it was also found that a majority of the malate flux is shuttled to the cytosol and plastid where it is converted to oxaloacetate providing reducing equivalents to these compartments. When compared to predictions by flux balance analysis, fluxes measured with (13)C-MFA were found to be suboptimal with respect to biomass yield; C. reinhardtii sacrifices biomass yield to produce ATP and reducing equivalents.