The Shewanella oneidensis MR - 1 fluxome under various oxygen conditions

The Shewanella oneidensis MR-1 fluxome under various oxygen conditions Abstract 1 The central metabolic fluxes of Shewanella oneidensis MR-1 were examined under 2 carbon-limited (aerobic) and oxygen-limited (micro-aerobic) chemostat conditions using 13 C 3 labeled lactate as the sole carbon source. The carbon labeling patterns of key amino acids in 4 biomass were probed using both GC-MS and 13 C-NMR. Based on the genome annotation, a 5 metabolic pathway model was constructed to quantify the central metabolic flux distributions. 6 The model showed that the tricarboxylic acid (TCA) cycle is the major carbon metabolism route 7 under both conditions. The Entner-Doudoroff and pentose phosphate pathways were utilized 8 primarily for biomass synthesis (flux below 5% of the lactate uptake rate). The anapleurotic 9 reactions (pyruvate to malate and oxaloacetate to phosphoenolpyruvate) and the glyoxylate shunt 10 were active. Under carbon-limited conditions, a substantial amount (9% of the lactate uptake 11 rate) of carbon entered the highly reversible serine metabolic pathway. Under micro-aerobic 12 conditions fluxes through the TCA cycle decreased and acetate production increased compared 13 to carbon-limited conditions, and flux from glyoxlate to glycine (serine-glyoxylate 14 aminotransferase) became measurable. Although flux distributions under aerobic, micro-aerobic, 15 and shake-flask culture conditions were different, the relative flux ratios of some central 16 metabolic reactions did not vary significantly (in particular, between shake flask and aerobic 17 chemostat). Hence, S. oneidensis central metabolism appears to be robust to environmental 18 changes. Our study also demonstrates the merit of coupling GC-MS with 13 C NMR for 19 metabolic flux analysis to reduce the use of 13 C labeled substrates and to obtain more accurate 20 flux values. 21