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Appl. Environ. Microbiol. doi:10.1128/AEM.02395-07
Copyright (c) 2008, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Codon-optimized bacterial genes improve L-arabinose fermentation in recombinant Saccharomyces cerevisiae

Beate Wiedemann and Eckhard Boles*

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

* To whom correspondence should be addressed. Email: e.boles{at}bio.uni-frankfurt.de.


  Abstract

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-ribluose-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 reaction 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.




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