Tuesday, April 30, 2013
Exhibit Hall
Metabolic profiling using nuclear magnetic resonance (NMR) spectroscopy enabled rapid and accurate identification and quantification of metabolites produced by the thermophilic obligately anaerobic cellulose-degrading gram positive bacterium Caldicellulosiruptor saccharolyticus, strain DSM 8903. NMR methods were used to characterize metabolites produced during growth on different monosaccharide carbon sources (D-glucose, D-mannose, L-arabinose, D-arabinose, D-xylose, L-fucose, and D-fucose). Several fermentation products whose formation would not have been predicted based on the current genome annotation were identified among the extracellular metabolites, indicating unexplored metabolic pathways in this organism. Data were collected on harvested culture aliquots, using culture supernatants to determine extracellular metabolites, and aqueous extracts of pelleted cells to determine intracellular metabolites. Analysis of 1-D 1H NMR spectra was accomplished by curve fitting against spectral libraries provided in Chenomx software. 2-D homonuclear and heteronuclear NMR experiments were employed to identify unassigned resonance peaks and to reduce uncertainties due to overlapping or poorly-resolved peaks. Additional confirmation of identifications for some metabolites was obtained by HPLC analysis. Novel fermentation products identified include: ethylene glycol (from growth on D-arabinose, though not L-arabinose), acetoin and 2,3-butanediol (from D-glucose and L-arabinose), and hydroxyacetone (from D-mannose and L-arabinose). Production of ethylene glycol from D-arabinose was particularly striking, with conversion of approximately 10% of the substrate carbon into this uncommon fermentation product. C. saccharolyticus has not previously been reported to produce these newly identified fermentation products, and these findings therefore suggest a previously unrealized metabolic potential for engineering C. saccharolyticus for synthesis of bioproducts.