ARS | Publication request: Cloning and Expression of Novel Bacterial Xylose Isomerases in Saccharomyces cerevisiae

Submitted to: Biotechnology for Fuels and Chemicals Symposium Proceedings
Publication Type: Abstract Only
Publication Acceptance Date: April 22, 2010
Publication Date: April 22, 2010
Citation: Hector, R.E., Dien, B.S., Cotta, M.A., Hughes, S.R. 2010. Cloning and expression of novel bacterial xylose isomerases in Saccharomyces cerevisiae. Biotechnology for Fuels and Chemicals Symposium. Abstract #1.

Technical Abstract: Saccharomyces yeasts are currently used for the industrial production of ethanol by fermentation of glucose derived from corn grain. These yeasts do not naturally consume the five-carbon sugars contained in the hemicellulose component of biomass feedstocks. Enzymes from native pentose-assimilating yeasts have been transferred to S. cerevisiae allowing fermentation of xylose, the most abundant biomass-derived pentose sugar. However, efficient conversion of xylose to ethanol is limited by cellular redox imbalance resulting from different co-factor specificities of the first two enzymes for xylose utilization (i.e., the xylose reductase and xylitol dehydrogenase enzymes). Replacing these two enzymes with a single, co-factor independent, xylose isomerase enzyme (XI) is proposed to alleviate this redox imbalance and improve xylose fermentation. Aside from an XI from the anaerobic fungus Piromyces, a very limited number of xylose isomerases have been demonstrated to function when expressed in S. cerevisiae. The goal of this study was to identify additional novel XI’s that function in S. cerevisiae. Xylose isomerase genes were isolated from multiple human and rumen intestinal bacteria by polymerase chain reaction (PCR) amplification with degenerate primers. These XI genes were expressed from plasmids in a S. cerevisiae strain that also expressed the S. cerevisiae xylulokinase gene at elevated levels. Xylose isomerase activities for these new enzymes and xylose fermentation data for the different XI-expressing S. cerevisiae strains will be presented.