Verlag des Forschungszentrums Jülich

JUEL-3538
Laufer, Birgit
Erweiterung des Substratspektrums von Zymomonas mobilis und Untersuchung xylose-verwertender Klone
99 S., 1998



The gram-negative bacterium Zymomonas mobilis ferments glucose, fructose and sucrose through the Entner-Doudoroff pathway efficiently to ethanol and CO₂. Attempts to broaden its narrow substrate range towards xylose have met for a time with limited success. Recombinant strains of Z. mobilis, which carried the genes for xylose metabolism (xylAB) and for transketolase (tktA) from Enterobacteria had not been able to grow on xylose as sole carbon source. The aim of the present work was the construction of recombinant strains of Zymomonas mobilis, expressing the plasmid-encoded genes xylAB and also the genes for transketolase (tktA) and transaldolase (talB) from Escherichia coli. It was conceived that these strains should able to ferment xylose efficiently to ethanol and CO₂. The recombinant strains expressed all necessary enzyme activities, but no immediate growth on xylose as sole carbon source was detected. In contrast, growth was inhibited when xylose was present as second carbon source. After a selection for growth in the presence of xylose, several clones were obtained which grew on xylose. It could be excluded that mutations in the plasmid-encoded genes enabled these clones to grow on xylose. However, the respective mutations in the chromosome could not been identified so far. The recombinant xylose-positive strain was cultured in batch fermentations with glucose (7%), xylose (7%), or glucose + xylose (2,5% each) as carbon source. Nearly all of the carbon was consumed and up to 96% of it was converted into ethanol and CO₂ as main products. Growth on xylose was slower and led to smaller growth yields than on glucose or glucose + xylose. Analysis of the by-products showed a transient appearance of xylulose in fermentations with xylose. In continuous fermentation with 1-¹³C-xylose, carbon flux in this recombinant strain was studied. The results, as well as the measurement of enzyme activities and the transient appearance of xylulose, indicated that xylulokinase is the limiting step in the xylose metabolism of the recombinant xylose positive strain.

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