Conversion of Mixed Sugars into Ethanol by Recombinant Corynebacterium Glutamicum

Masayuki Inui, Hideo Kawaguchi, Shohei Okino, Masako Suda, Miho Sasaki, Alain A. Vertès, and Hideaki Yukawa. Microbiology Research Group, Research Institute of Innovative Technology for the Earth (RITE), 9-2 Kizugawadai, Kizu-cho, Soraku-gun, Kyoto, Japan

We conducted whole-cell catalysis using growth-arrested Corynebacterium glutamicum to produce ethanol from pentose sugars. This recombinant catalysis process was enabled by the introduction into the C. glutamicum genome of various genes, including particularly the pyruvate dehydrogenase and lactate dehydrogenase encoding genes from Zymomonas mobilis 1), and the xylose isomerase and xylose reductase encoding genes of Escherichia coli 2). Furthermore, we combined process engineering and metabolic engineering techniques to optimize ethanol production by suppressing the secretion of various fermentation by-products such as lactate, acetate, and succinate.

The recombinant strain thus created was used in a growth-arrested process where product production phase and cell catalyst production phase are uncoupled. Growth-cessation is achieved simply by the lowering of the oxygen tension which results when a high density cell suspension is incubated in the absence of aeration. The main metabolic machinery remains however functional and the production reaction can be continued for several weeks using the same growth-arrested cells. An important property of this design is that the bulk of the carbon source is directed towards product production rather than towards vegetative functions. Ethanol productivities greater than 30 g/l/h could be routinely attained 1).

This work paves the way for the construction of an efficient bioconverter strain capable of cost-effectively utilising various sugars, and particularly the pentose sugars that are typically contained in saccharified lignocellulosic mixtures. Notably, the fast on/off response displayed by the process/converter strain system that we propose facilitates the implementation of continuous ethanol fermentation production that can be run at steady-state for an extended period of time. This property is critical for the realization of the biorefinery concept.

This study was partially supported by a grant from New Energy and Industrial Technology Development Organization (NEDO).

1)M. Inui, H. Kawaguchi, S. Murakami, A.A. Vertès and H. Yukawa. Metabolic engineering of Corynebacterium glutamicum for fuel ethanol production under oxygen-deprivation conditions. J. Mol. Microbiol. Biotechnol. 8:243-254. 2004. 2)H. Kawaguchi, A.A. Vertès, S. Okino, M. Inui and H. Yukawa. Engineering of a xylose metabolic pathway in Corynebacterium glutamicum. Appl. Environ. Microbiol. 72:3418-3428. 2006.

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