ORNL researchers develop acetate-tolerant microorganism

By Lisa Gibson
Posted June 2, 2010, at 9:20 a.m. CST

Scientists from the U.S. DOE's Oak Ridge National Laboratory say they have developed a microorganism with an improved ability to convert wood products to biofuel. The researchers identified a key Zymomonas mobilis gene that, when over expressed, allows the microorganism to resist inhibitors.

Currently, biomass materials such as corn stover and switchgrass must undergo pretreatments to loosen the cellular structure enough to extract the sugar from the cellulose. Those treatments add new challenges because they create chemicals called inhibitors that stop microorganisms from performing fermentation. There are two ways to combat that problem, according to Steven Brown, researcher in ORNL's Bioenergy Research Center. "One way is to remove the inhibitors, but this method is very expensive and would not help biofuels become cost-competitive with gasoline," he said. "The second way is what we do, which is to develop microorganisms that are more tolerant of the inhibitors."

The nonmutated strain of Z. mobilis cannot grow in the presence of acetate, a predominant inhibitor. But when a slice of DNA containing the gene NhaA is inserted into the nonmutated strain, the bacterium can withstand acetate in its environment, according to ORNL. "We identified a deletion in the genome of the mutant strain that allowed much NhaA to be made in the mutant relative to the wild-type strain," Brown said. "The over expression of this gene, which codes for a sodium-proton antiporter, allows the bacterium to grow in concentrations of sodium acetate that the wild type cannot stand."

The research team looked at related genes in other microorganisms and found that the method translates. It is unclear why the NhaA gene affects Z. mobilis's immunity to acetate and more research in the area needs to be done, Brown said, but related bacterium Escherichia coli uses the protein to pump sodium ions out of the cell while importing protons. "We have conducted some structural studies on the Z. mobilis NhaA protein and it appears to be similar to E. coli protein, but the exact nature of sodium ion transport and what happens to the acetate remain to be determined in Z. mobilis," he said.

Brown and his colleague Shihui Yang have authored papers on their work for the online Proceedings of the National Academy of Sciences and BMC Microbiology. They have examined model inhibitor compounds to date, but there are likely to be synergistic effects in real biomass materials, Brown said. "It would be very interesting to test the strains on a pretreated biomass material, such as corn stover or switchgrass, and ideal if a company found our discoveries useful," he added. "They are available for license and perhaps they might allow a company to make more rapid gains for strain development."

The team is examining other inhibitors and has recently filed a provisional patent application on a single mutated gene that confers substantially greater ethanol tolerance, Brown said. "This is a particularly important area to make progress in," he said, adding the novel finding has profound implications for the field.

"I hope that other researchers, economists and scientists will examine our study and find utility that could bring down process costs," Brown said. "We are conducting basic research funded by the (U.S. DOE) Office of Science and ultimately the promise of the research will be determined by others in the field and industry."