Researchers at Georgia Tech have applied adaption (adaptive mutation) to a strain of the bacterium Zymomonas mobilis to increase its ability to produce ethanol. The research, published in the journal Biotechnology and Bioengineering, shows how adaptation and metabolic engineering can be combined for strain improvement.
Z. mobilis is noted for its bio-ethanol producing potential, with productivity exceeding yeast strains by several fold. Metabolic engineering has expanded its substrate range to include xylose; however, xylose fermentation has lagged far behind glucose. In addition, xylose fermentation is also incomplete. The team believed that ethanol production could be increased through improvement of xylose fermentation.
In this study we sought to improve ethanol production by enhancing the ability of Z. mobilis to use and ferment xylose. Fermenting xylose at high concentration could in turn increase ethanol concentration, resulting in much improved productivity.—Dr. Rachel Chen
As a result of adaptation over 80 days and 30 serial transfers in a medium containing high concentration of xylose, the team obtained a strain, referred as A3, with markedly improved xylose metabolism. The new strain was able to grow on 10% (w/v) xylose and rapidly ferment xylose to ethanol within 2 days and retained high ethanol yield.
Similarly, in mixed glucose-xylose fermentation, a total of 9% (w/v) ethanol—the highest ever shown for this organism in mixed sugar fermentation, according to Chen—was obtained from two doses of 5% glucose and 5% xylose (or a total of 10% glucose and 10% xylose).
The team found that by altering the strain, sugar fermentation time was reduced from over 110 hours to about 35 hours. This improvement in fermentation allowed the strain to ferment higher concentrations of xylose.
This research also investigated the underlying mechanism for the improvement. Interestingly, by adapting a strain in a high concentration of xylose, significant alterations of metabolism occurred.
One noticeable change was reduced levels of xylitol, a byproduct of xylose fermentation which can inhibit the strain’s xylose metabolism. In addition, the first step of xylose metabolism, believed to be the rate-limiting step, was accelerated 4-8 times in the adapted strain, with the net effect of channeling xylose to ethanol instead of xylitol.
This research illustrates the power of adaptation in strain improvement. This confirms that xylitol metabolism is the key to efficient use of xylose in this bacterium, which in turn can be vital for producing ethanol. This shows that adaptation is not only useful in improving strains, but is equally useful for pinpointing key bottlenecks in metabolically engineered strains.—Rachel Chen
Agrawal. M, Mao. Z, Chen. R (2010) Adaptation yields a highly efficient xylose-fermenting Zymomonas mobilis strain. Biotechnology and Bioengineering, doi: 10.1002/bit.23021