Metabolic state of Zymomonas mobilis in glucose-, fructose-, and xylose-fed continuous cultures as analysed by C-13- and P-31-NMR spectroscopy

The reasons for the well-known significantly different behaviour of the ana erobic, gram-negative, ethanologenic bacterium Zymomonas mobilis during gro wth on fructose (i.e. decreased growth and ethanol yields, increased by-pro duct formation) as compared to that on its second natural substrate, glucos e, have remained unexplained. A xylose-fermenting recombinant strain of Z. mobilis that was recently constructed in our laboratory also unexpectedly d isplayed an increased formation of byproducts and a strongly reduced growth rate as compared to the parent strain. Therefore, a comprehensive study em ploying recently developed NMR-based methods for the in vivo analysis of in tracellular phosphorylated pool sizes and metabolic fluxes was undertaken t o enable a global characterization of the intracellular metabolic state of Z. mobilis during growth on C-13-labelled glucose, fructose and xylose in d efined continuous cultures. The C-13-NMR flux analysis indicated that ribos e 5-phosphate is synthesized via the nonoxidative pentose phosphate pathway in Z. mobilis, and it identified a metabolic bottleneck in the recombinant xylose-fermenting Z. mobilis strain at the level of heterologous xylulokin ase, The P-31-NMR analyses revealed a global alteration of the levels of in tracellular phosphorylated metabolites during growth on fructose as compare d to that on glucose. The results suggest that this is primarily caused by an elevated concentration of intracellular fructose 6-phosphate.