Saccharomyces cerevisiae is considered one of the most promising organisms for ethanol production from lignocellulosic feedstock. Unfortunately, pentose sugars, which comprise up to 30% of lignocellulose, cannot be utilized by wild type S. cerevisiae. Heterologous pathways were introduced into S. cerevisiae to enable utilization of d-xylose, the most abundant pentose sugar. However, the resulting recombinant S. cerevisiae strains exhibited a slow growth rate and poor sugar utilization efficiency when grown on d-xylose as the sole carbon source. d-xylose uptake is the first step of d-xylose utilization. d-xylose can only enter yeast cells through hexose transporters, which have two orders of magnitude lower affinity towards d-xylose compared to hexoses. It was also shown that inefficient pentose uptake is the limiting step in some d-xylose metabolizing yeast strains. Here we report the cloning and characterization of two novel d-xylose-specific transporters from Neurospora crassa and Pichia stipitis. These two transporters were identified from a total of 18 putative pentose transporters. They were functionally expressed and properly localized in S. cerevisiae as indicated by HPLC analysis and fluorescence confocal microscopy, respectively. Kinetic parameters of the d-xylose-specific transporters were determined using a 14C-labeled sugar uptake assay. Use of pentose-specific transporters should improve d-xylose consumption and ethanol production in fast d-xylose assimilating strains, thereby lowering the cost of lignocellulosic ethanol production.
ASJC Scopus subject areas
- Molecular Biology