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Codon-Optimized Bacterial Genes Improve L-Arabinose Fermentation in Recombinant Saccharomyces cerevisiae

Institute of Molecular Biosciences, Goethe-Universität Frankfurt am Main, Max-von-Laue-Str. 9, D-60438 Frankfurt am Main, Germany

Received 24 October 2007/ Accepted 2 February 2008

Bioethanol produced by microbial fermentations of plant biomass hydrolysates consisting of hexose and pentose mixtures is an excellent alternative to fossil transportation fuels. However, the yeast Saccharomyces cerevisiae, commonly used in bioethanol production, can utilize pentose sugars like L-arabinose or D-xylose only after heterologous expression of corresponding metabolic pathways from other organisms. Here we report the improvement of a bacterial L-arabinose utilization pathway consisting of L-arabinose isomerase from Bacillus subtilis and L-ribulokinase and L-ribulose-5-P 4-epimerase from Escherichia coli after expression of the corresponding genes in S. cerevisiae. L-Arabinose isomerase from B. subtilis turned out to be the limiting step for growth on L-arabinose as the sole carbon source. The corresponding enzyme could be effectively replaced by the enzyme from Bacillus licheniformis, leading to a considerably decreased lag phase. Subsequently, the codon usage of all the genes involved in the L-arabinose pathway was adapted to that of the highly expressed genes encoding glycolytic enzymes in S. cerevisiae. Yeast transformants expressing the codon-optimized genes showed strongly improved L-arabinose conversion rates. With this rational approach, the ethanol production rate from L-arabinose could be increased more than 2.5-fold from 0.014 g ethanol h^–1 (g dry weight)^–1 to 0.036 g ethanol h^–1 (g dry weight)^–1 and the ethanol yield could be increased from 0.24 g ethanol (g consumed L-arabinose)^–1 to 0.39 g ethanol (g consumed L-arabinose)^–1. These improvements make up a new starting point for the construction of more-efficient industrial L-arabinose-fermenting yeast strains by evolutionary engineering.

Published ahead of print on 8 February 2008.