C. autoethanogenum is able to co-produce 2,3-butanediol in significant amounts
along with the major product ethanol (with a ratio of 1:10) from CO-containing
steel mill waste gas [81]. 2,3-BD is an important high-valuable chemical precursor
for production of a variety of compounds such as methyl ethyl ketone (MEK) which
is used in printing inks as well as in surface coatings, and 1,3-butadiene, an
intermediate used for production of nylon and synthetic rubber [84, 85]. The main
2,3-BD derived products have an immense global market and are, therefore, of high
economic interest.
In C. autoethanogenum, the production of 2,3-BD starts with the conversion of
acetyl-CoA to pyruvate catalyzed by the PFOR. C. autoethanogenum possesses two
identical genes encoding for the PFOR. In a next step, two molecules of pyruvate
are then converted to acetolactate by an acetolactate synthase. C. autoethanogenum
possesses three different genes encoding for acetolactate synthases [81]. The genes
ilvB and ilvIH encoding for two of the three acetolactate synthases are most
probably involved in the anabolism, whereas the gene alsS encodes most likely for
the acetolactate synthase responsible for the catabolic formation of 2,3-butanediol.
The formed acetolactate is then further split into acetoin and CO 2 by an acetolactate
decarboxylase (ALDC). In a final step, acetoin is reduced to 2,3-butanediol by
either a NADH-dependent 2,3-butanediol dehydrogenase or a NADPH-dependent
primary-secondary alcohol dehydrogenase.
Even though C. autoethanogenum can naturally produce high amounts of
ethanol and also significant amounts of 2,3-BD under autotrophic conditions, a
problem that comes along with the industrial scaled production of both products is
an undesired production of unwanted side-products such as acetate or lactate (in
minor amounts). Especially, the production of lactate as side-product significantly
reduces efficiency and yield of the produced 2,3-BD/ethanol. Furthermore, low
concentrations of lactate (<0.5%) are toxic to C. autoethanogenum [81]. Another
problem is that lactate can also be used as substrate by other microorganisms which
increases the chances of a contamination during the fermentation process when
lactate is formed. A big advantage to overcome these problems is the genetical
accessibility of C. autoethanogenum, making metabolic engineering possible.
Strategies to improve ethanol production on a genetic level are mainly based on
either overexpression or on introduction of AdhE. Another promising approach to
enhance yields and production rates by metabolic engineering includes the deletion
of genes via allelic exchange mutagenesis [86]. By deletion of the gene encoding
for the lactate dehydrogenase in C. autoethanogenum, the production of 2,3-BD
and ethanol was improved [87]. With the same genetic tool, the genes adhE and
aor2 were deleted which also resulted in an improved ethanol production (up to
180% increase) under autotrophic conditions [88].
3.4 Clostridium carboxidivorans
The acetogenic bacterium C. carboxidivorans is known to produce several solvents
such as ethanol, butanol, hexanoic acid and hexanol from syngas [89]. It was
Acetogenic Bacteria for Biotechnological Applications
119
along with the major product ethanol (with a ratio of 1:10) from CO-containing
steel mill waste gas [81]. 2,3-BD is an important high-valuable chemical precursor
for production of a variety of compounds such as methyl ethyl ketone (MEK) which
is used in printing inks as well as in surface coatings, and 1,3-butadiene, an
intermediate used for production of nylon and synthetic rubber [84, 85]. The main
2,3-BD derived products have an immense global market and are, therefore, of high
economic interest.
In C. autoethanogenum, the production of 2,3-BD starts with the conversion of
acetyl-CoA to pyruvate catalyzed by the PFOR. C. autoethanogenum possesses two
identical genes encoding for the PFOR. In a next step, two molecules of pyruvate
are then converted to acetolactate by an acetolactate synthase. C. autoethanogenum
possesses three different genes encoding for acetolactate synthases [81]. The genes
ilvB and ilvIH encoding for two of the three acetolactate synthases are most
probably involved in the anabolism, whereas the gene alsS encodes most likely for
the acetolactate synthase responsible for the catabolic formation of 2,3-butanediol.
The formed acetolactate is then further split into acetoin and CO 2 by an acetolactate
decarboxylase (ALDC). In a final step, acetoin is reduced to 2,3-butanediol by
either a NADH-dependent 2,3-butanediol dehydrogenase or a NADPH-dependent
primary-secondary alcohol dehydrogenase.
Even though C. autoethanogenum can naturally produce high amounts of
ethanol and also significant amounts of 2,3-BD under autotrophic conditions, a
problem that comes along with the industrial scaled production of both products is
an undesired production of unwanted side-products such as acetate or lactate (in
minor amounts). Especially, the production of lactate as side-product significantly
reduces efficiency and yield of the produced 2,3-BD/ethanol. Furthermore, low
concentrations of lactate (<0.5%) are toxic to C. autoethanogenum [81]. Another
problem is that lactate can also be used as substrate by other microorganisms which
increases the chances of a contamination during the fermentation process when
lactate is formed. A big advantage to overcome these problems is the genetical
accessibility of C. autoethanogenum, making metabolic engineering possible.
Strategies to improve ethanol production on a genetic level are mainly based on
either overexpression or on introduction of AdhE. Another promising approach to
enhance yields and production rates by metabolic engineering includes the deletion
of genes via allelic exchange mutagenesis [86]. By deletion of the gene encoding
for the lactate dehydrogenase in C. autoethanogenum, the production of 2,3-BD
and ethanol was improved [87]. With the same genetic tool, the genes adhE and
aor2 were deleted which also resulted in an improved ethanol production (up to
180% increase) under autotrophic conditions [88].
3.4 Clostridium carboxidivorans
The acetogenic bacterium C. carboxidivorans is known to produce several solvents
such as ethanol, butanol, hexanoic acid and hexanol from syngas [89]. It was
Acetogenic Bacteria for Biotechnological Applications
119