Evaluation of Enzyme Immobilization Methods for Xylose-to-Xylulose Isomerization

Bioconversion of lignocellulosic feedstocks to fuel-grade ethanol is a promising route towards reduced dependence on fossil fuels and the mitigation of greenhouse gas production. Baker's yeast (Saccharomyces cerevisiae) used for industrial-scale ethanol production is unable to utilize xylose, which accounts for about 30% of the total sugars released in biomass hydrolysate. Current methods to overcome this limitation are focused on development of different types of genetically modified organisms (GMOs). However, several problems exist for these GMOs. First, the consumption rate of xylose is low, and the time required to produce ethanol is relatively long. This low rate of xylose fermentation is related to the intracellular conversion of xylose to xylulose which has an unfavorable equilibrium. In addition, the genetic stability of GMOs, costs for propagation, and robustness under industrial fermentation conditions has not been reported.

In our approach, we propose using immobilized xylose isomerase (XI) for the extracellular conversion of xylose to xylulose, followed by fermentation of xylulose to ethanol by native yeast. To implement the use of immobilized XI under conditions suitable for fermentation, we co-immobilize XI with urease. We have already demonstrated proof-of-principle for the co-immobilized enzymes' effectiveness at isomerizing xylose at low pH. Thus, for cost-effective use of these co-immobilized enzymes, methods must be developed for robust immobilization of the enzymes as well as for ease of reuse and recovery of the enzyme pellets. In this poster, we present the results of several methods we have tested for the immobilization of the XI enzyme. Pellet lifetimes as well as pH and temperature behavior of the immobilized XI is presented.