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Chapter 3 : Engineering the Genome for Fermentation of Hemicellulose Hydrolysates

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Engineering the Genome for Fermentation of Hemicellulose Hydrolysates, Page 1 of 2

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Abstract:

is a source of genes for engineering xylose metabolism in task undertaken in numerous laboratories around the world. Increasing the capacity of for rapid xylose fermentation can greatly improve its usefulness in commercial xylose fermentations. Genetic transformation with is more efficient than transformation with the modified marker. Cultivation conditions can strongly affect expression of fermentative enzymes in . Unlike , which regulates fermentation by sensing the presence of glucose, induces fermentative activity in response to oxygen limitation. The coupling of Xyl1 and Xyl2 activities therefore tends to result in the consumption of NADPH and accumulation of NADH. Preliminary data based on expressed sequence tags indicate that transcripts for fatty acid synthase (FAS2) stearoyl-coenzyme A desaturase (OLE1) are induced under oxygen-limiting conditions. Xylanase production by has been recognized, and the organism has also been transformed with heterologous xylanases to increase xylanase activity. A genetic system has been developed that includes the auxotrophic markers URA3 and LEU3 along with modified forms of the phleomycin D1 resistance marker, , and the recombinase. Many genes in are found in functionally related clusters. Further metabolic engineering and strain selection are needed to increase the overall fermentation rate and ethanol tolerance of for the commercial bioconversion of hemicellulose hydrolysates.

Citation: Jeffries T. 2008. Engineering the Genome for Fermentation of Hemicellulose Hydrolysates, p 37-47. In Wall J, Harwood C, Demain A (ed), Bioenergy. ASM Press, Washington, DC. doi: 10.1128/9781555815547.ch3

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Figures

Image of Figure 1.
Figure 1.

Mutants developed from CBS 6054.

Citation: Jeffries T. 2008. Engineering the Genome for Fermentation of Hemicellulose Hydrolysates, p 37-47. In Wall J, Harwood C, Demain A (ed), Bioenergy. ASM Press, Washington, DC. doi: 10.1128/9781555815547.ch3
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Image of Figure 2.
Figure 2.

Relative expression of transcripts for the glutamate decarboxylase bypass. Abbreviations: and , isocitrate dehydrogenase 1 and 2; , NAD-specific glutamate dehydrogenase; , NADP-specific glutamate dehydrogenase;, glutamate decarboxylase 2; and , 4-aminobutyrate aminotransferase; and , succinate semialdehyde dehydrogenase; , 2-ketoglutarate dehydrogenase; Isoct, isocitrate; AKG, 2-keto-glutarate; LGlu, L-glutamate; 4-AB, 4-aminobutyrate; Suc-SA, succinate semialdehyde; Succ, succinate; Fum, fumarate; GA, glucose aerobic; XA, xylose aerobic; GOL, glucose oxygen limited; XOL, xylose oxygen limited.

Citation: Jeffries T. 2008. Engineering the Genome for Fermentation of Hemicellulose Hydrolysates, p 37-47. In Wall J, Harwood C, Demain A (ed), Bioenergy. ASM Press, Washington, DC. doi: 10.1128/9781555815547.ch3
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Image of Figure 3.
Figure 3.

Induction of transcripts for lipid synthesis under oxygen-limiting conditions. Abbreviations: GA, glucose aerobic; XA, xylose aerobic; GOL, glucose oxygen limited; XOL, xylose oxygen limited.

Citation: Jeffries T. 2008. Engineering the Genome for Fermentation of Hemicellulose Hydrolysates, p 37-47. In Wall J, Harwood C, Demain A (ed), Bioenergy. ASM Press, Washington, DC. doi: 10.1128/9781555815547.ch3
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Image of Figure 4.
Figure 4.

Gene clusters of β-glucosidases and endoglucanases with sugar transporters in the genome. Approximate chromosome coordinates are shown. All constructs were derived from the original sequence deposit at the Joint Genome Institute website (http://genome.jgi-psf.org/Picst3/Picst3.home.html). The complete genome is also found in GenBank.

Citation: Jeffries T. 2008. Engineering the Genome for Fermentation of Hemicellulose Hydrolysates, p 37-47. In Wall J, Harwood C, Demain A (ed), Bioenergy. ASM Press, Washington, DC. doi: 10.1128/9781555815547.ch3
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Image of Figure 5.
Figure 5.

Phylogenetic relationships among hexose transporters and beta-glucosidases found in clusters in . Gene names correspond to designations given on the Joint Genome Institute website (see legend to Fig. 4 for URL) and in GenBank. The method used was Neighbor Joining with Best Tree; distances were uncorrected, and gaps were distributed proportionally.

Citation: Jeffries T. 2008. Engineering the Genome for Fermentation of Hemicellulose Hydrolysates, p 37-47. In Wall J, Harwood C, Demain A (ed), Bioenergy. ASM Press, Washington, DC. doi: 10.1128/9781555815547.ch3
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Image of Figure 6.
Figure 6.

Gene clusters of α-glucosidases with putative maltose permeases in the genome. Approximate chromo-some coordinates are shown. All constructs were derived from the original sequence deposit at the Joint Genome Institute website (see legend to Fig. 4 for URL). The complete genome is also found in GenBank.

Citation: Jeffries T. 2008. Engineering the Genome for Fermentation of Hemicellulose Hydrolysates, p 37-47. In Wall J, Harwood C, Demain A (ed), Bioenergy. ASM Press, Washington, DC. doi: 10.1128/9781555815547.ch3
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Image of Figure 7.
Figure 7.

Phylogenetic relationships of putative sugar transporters from . Except as noted, all genes are from . Gene names correspond to designations given on the Joint Genome Institute website and in GenBank. Sequences for and are from ; sequences for and homolog are from . The method used was Neighbor Joining with Best Tree; distances were uncorrected, and gaps were distributed proportionally.

Citation: Jeffries T. 2008. Engineering the Genome for Fermentation of Hemicellulose Hydrolysates, p 37-47. In Wall J, Harwood C, Demain A (ed), Bioenergy. ASM Press, Washington, DC. doi: 10.1128/9781555815547.ch3
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Image of Figure 8.
Figure 8.

Phylogenetic relationships of α-glucosidases from . Gene names correspond to designations given on the Joint Genome Institute website and in GenBank. The method used was Neighbor Joining with Best Tree; distances were uncorrected, and gaps were distributed proportionally.

Citation: Jeffries T. 2008. Engineering the Genome for Fermentation of Hemicellulose Hydrolysates, p 37-47. In Wall J, Harwood C, Demain A (ed), Bioenergy. ASM Press, Washington, DC. doi: 10.1128/9781555815547.ch3
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Image of Figure 9.
Figure 9.

Galactose and lactose gene clusters in chromosome 3.

Citation: Jeffries T. 2008. Engineering the Genome for Fermentation of Hemicellulose Hydrolysates, p 37-47. In Wall J, Harwood C, Demain A (ed), Bioenergy. ASM Press, Washington, DC. doi: 10.1128/9781555815547.ch3
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Tables

Generic image for table
Table 1.

Fermentation characteristics of three strains of grown in shake flasks

Citation: Jeffries T. 2008. Engineering the Genome for Fermentation of Hemicellulose Hydrolysates, p 37-47. In Wall J, Harwood C, Demain A (ed), Bioenergy. ASM Press, Washington, DC. doi: 10.1128/9781555815547.ch3

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