Second-Generation Bioethanol: Advancement of Ethanologenic …
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medium (Matsushika et al. 2009). P. stipitis is one of the few yeasts that possess the
ability to excrete less xylitol than other yeasts (Debus et al. 1983).
Theoretically, prokaryotic XI should be more efficient in metabolizing xylose
compared to yeasts because no cofactor is necessary to produce D-xylulose in
prokaryote hence intracellular redox imbalance will not occur. Few studies have
been conducted which involve two strategies, the expression of prokaryotic XI in
high yield ethanologenic yeast and the simultaneous isomerization and fermentation
of D-xylose to ethanol. In the former, Brat et al. (2009) have accomplished heterologous expression of XI from bacteria, Clostridium phytofermentansthat have enabled
S. cerevisiae to metabolize xylose. However, the growth rates of their recombinant
strains are rather low with ethanol yield at 0.43 g ethanol g D-xylose
−1 . While in the
latter, two-step process by Gong et al. (1981) allows non-xylose-fermenting yeast to
ferment xylose using preceding prokaryotic XI in a separate process. The separate
isomerization is preferable due to different optimal conditions for isomerization and
fermentation. The results indicate that ethanol could be produced from D-xylose with
a yield of greater than 80%.
The latest discovery of xylose metabolic pathway was in halophilic archaea such
as Haloferax volcanii and Haloarcula marismortui (Johnsen and Schönheit 2004;
Johnsen et al. 2009). The catabolism occurs through the oxidation of D-xylose to an
intermediate of tricarboxylic acid cycle, α-ketoglutarate. The enzymes involved in the
degradation process are D-xylose dehydrogenase, xylonate dehydratase, 2-keto-3deoxyxylonate dehydratase, and α-ketoglutarate semialdehyde dehydrogenase. This
finding definitely opens a new opportunity toward further manipulation of xylose
metabolic pathway. However, due to the inclusion of multiple genes and conversion
complexity, there is no research on expressing this metabolic pathway on potential
ethanologens yet.
4 On Improving Tolerance Toward Inhibitors
The utilization of lignocellulosic biomass demands advantageous traits from the
potential ethanologens. These are due to difficulties in achieving an efficient model
of integrated process involving the selection of biomass toward the harvesting of
bioethanol. Prior to fermentation, pretreatment and hydrolysis processes generate
few inhibitors commonly lignin residues, acids, and aldehydes. To date, there was
no known method to avoid the formation of these inhibitors except for less severe
pretreatment and detoxification process for reducing inhibitors content (Palmqvist
and Hahn-Hägerdal 2000). However, one of the drawbacks for such process is the
extra cost, which burden on attempts to market bioethanol at a low price (Taylor et al.
2012). Thus, an apparent solution is to enhance the ability of potential ethanologens
to become tolerant to these inhibitors.
71
medium (Matsushika et al. 2009). P. stipitis is one of the few yeasts that possess the
ability to excrete less xylitol than other yeasts (Debus et al. 1983).
Theoretically, prokaryotic XI should be more efficient in metabolizing xylose
compared to yeasts because no cofactor is necessary to produce D-xylulose in
prokaryote hence intracellular redox imbalance will not occur. Few studies have
been conducted which involve two strategies, the expression of prokaryotic XI in
high yield ethanologenic yeast and the simultaneous isomerization and fermentation
of D-xylose to ethanol. In the former, Brat et al. (2009) have accomplished heterologous expression of XI from bacteria, Clostridium phytofermentansthat have enabled
S. cerevisiae to metabolize xylose. However, the growth rates of their recombinant
strains are rather low with ethanol yield at 0.43 g ethanol g D-xylose
−1 . While in the
latter, two-step process by Gong et al. (1981) allows non-xylose-fermenting yeast to
ferment xylose using preceding prokaryotic XI in a separate process. The separate
isomerization is preferable due to different optimal conditions for isomerization and
fermentation. The results indicate that ethanol could be produced from D-xylose with
a yield of greater than 80%.
The latest discovery of xylose metabolic pathway was in halophilic archaea such
as Haloferax volcanii and Haloarcula marismortui (Johnsen and Schönheit 2004;
Johnsen et al. 2009). The catabolism occurs through the oxidation of D-xylose to an
intermediate of tricarboxylic acid cycle, α-ketoglutarate. The enzymes involved in the
degradation process are D-xylose dehydrogenase, xylonate dehydratase, 2-keto-3deoxyxylonate dehydratase, and α-ketoglutarate semialdehyde dehydrogenase. This
finding definitely opens a new opportunity toward further manipulation of xylose
metabolic pathway. However, due to the inclusion of multiple genes and conversion
complexity, there is no research on expressing this metabolic pathway on potential
ethanologens yet.
4 On Improving Tolerance Toward Inhibitors
The utilization of lignocellulosic biomass demands advantageous traits from the
potential ethanologens. These are due to difficulties in achieving an efficient model
of integrated process involving the selection of biomass toward the harvesting of
bioethanol. Prior to fermentation, pretreatment and hydrolysis processes generate
few inhibitors commonly lignin residues, acids, and aldehydes. To date, there was
no known method to avoid the formation of these inhibitors except for less severe
pretreatment and detoxification process for reducing inhibitors content (Palmqvist
and Hahn-Hägerdal 2000). However, one of the drawbacks for such process is the
extra cost, which burden on attempts to market bioethanol at a low price (Taylor et al.
2012). Thus, an apparent solution is to enhance the ability of potential ethanologens
to become tolerant to these inhibitors.