highest final acetate concentration reported so far could be achieved by using a
pH-controlled, batch-operated stirred-tank bioreactor. However, problems that have
to be overcome to use A. woodii as industrial platform with autotrophic conditions
are a rather low solubility of the gaseous substrate and poor growth of biomass
under these conditions [46].
A special feature of A. woodii is the presence of an enzyme complex that
catalyzes the first step in the WLP, the reduction of CO 2 to formic acid. Typically,
this reaction is catalyzed by a NADP- or ferredoxin-dependent formate dehydrogenases [19–21], whereas in A. woodii, as well as in T. kivui, this reaction is
catalyzed by a novel enzyme, a hydrogen-dependent CO 2 reductase (HDCR) [14,
15]. The HDCR is composed of four subunits. Both HDCRs consist of a formate
dehydrogenase module and a [FeFe] hydrogenase module (HydA2) that are connected by two small FeS-containing subunits which likely transfer electrons from
the hydrogenase module to the formate dehydrogenase module and vice versa.
A. woodii has two isoforms of the HDCR: one containing a cysteine and the other a
selenocysteine. T. kivui only has one isoenzyme with a cysteine in the formate
dehydrogenase module. In contrast to formate dehydrogenases, the HDCR directly
catalyzes the reduction of CO 2 to formic acid with H 2 as electron donor with rates
superior to any known chemical catalyst, thus, making it a promising candidate for
storage of H 2 [14, 15]. Molecular hydrogen is an attractive future energy carrier to
replace fossil fuels. The limiting problem for the technological use of H 2 is storage
of the gas. However, this problem could be overcome by using the HDCR. Since
the equilibrium constant for the hydrogenation of CO 2 to formate is close to one,
the HDCR is an ideal biocatalyst for the storage of molecular hydrogen. In the form
of formate, the so-called liquid organic hydrogen carrier (LOHC), the explosive H 2
could easily be handled and supplied by using existing infrastructure [49, 50]. The
HDCRs from both acetogens are significantly more effective than the best chemical
catalysts known so far [49, 51–53]. The HDCR from A. woodii catalyzes the
hydrogenation of CO 2 with a turnover frequency (TOF) of 101 600 h
−1 at 30 °C
and 0.8 bar H 2 , being significantly better than chemical catalysts. Even though the
enzyme of A. woodii is highly stable with a temperature optimum of 40 °C,
long-term stability has yet to be determined. Remarkably, the [FeFe] hydrogenase
module of the HDCR is the [FeFe] hydrogenase being completely CO tolerant [54].
In contrast to other [FeFe] hydrogenases where CO leads to irreversible damage to
the H-cluster [55–57], CO inhibition of the [FeFe] hydrogenase module is fully
reversible, even though the enzyme is strongly inhibited by CO [54]. This feature
could be a result of the metabolism of A. woodii, where CO is an intermediate of the
WLP. However, since the HDCR requires strict anoxic conditions and/or a low
redox potential, a biotechnological application is rather difficult. To overcome this
problem, we established a whole-cell system for the efficient hydrogenation of CO 2
to formate [15]. The system is based on the knockout of the ATP-dependent further
conversion of formate in the WLP of acetogens. Formate is transiently excreted and
accumulated during autotrophic growth in A. woodii [58] and accumulation requires
inhibition of the ATP-dependent further conversion of formate in the WLP. By
reducing the cellular energy content, formate is no longer converted to acetate and
Acetogenic Bacteria for Biotechnological Applications
115
pH-controlled, batch-operated stirred-tank bioreactor. However, problems that have
to be overcome to use A. woodii as industrial platform with autotrophic conditions
are a rather low solubility of the gaseous substrate and poor growth of biomass
under these conditions [46].
A special feature of A. woodii is the presence of an enzyme complex that
catalyzes the first step in the WLP, the reduction of CO 2 to formic acid. Typically,
this reaction is catalyzed by a NADP- or ferredoxin-dependent formate dehydrogenases [19–21], whereas in A. woodii, as well as in T. kivui, this reaction is
catalyzed by a novel enzyme, a hydrogen-dependent CO 2 reductase (HDCR) [14,
15]. The HDCR is composed of four subunits. Both HDCRs consist of a formate
dehydrogenase module and a [FeFe] hydrogenase module (HydA2) that are connected by two small FeS-containing subunits which likely transfer electrons from
the hydrogenase module to the formate dehydrogenase module and vice versa.
A. woodii has two isoforms of the HDCR: one containing a cysteine and the other a
selenocysteine. T. kivui only has one isoenzyme with a cysteine in the formate
dehydrogenase module. In contrast to formate dehydrogenases, the HDCR directly
catalyzes the reduction of CO 2 to formic acid with H 2 as electron donor with rates
superior to any known chemical catalyst, thus, making it a promising candidate for
storage of H 2 [14, 15]. Molecular hydrogen is an attractive future energy carrier to
replace fossil fuels. The limiting problem for the technological use of H 2 is storage
of the gas. However, this problem could be overcome by using the HDCR. Since
the equilibrium constant for the hydrogenation of CO 2 to formate is close to one,
the HDCR is an ideal biocatalyst for the storage of molecular hydrogen. In the form
of formate, the so-called liquid organic hydrogen carrier (LOHC), the explosive H 2
could easily be handled and supplied by using existing infrastructure [49, 50]. The
HDCRs from both acetogens are significantly more effective than the best chemical
catalysts known so far [49, 51–53]. The HDCR from A. woodii catalyzes the
hydrogenation of CO 2 with a turnover frequency (TOF) of 101 600 h
−1 at 30 °C
and 0.8 bar H 2 , being significantly better than chemical catalysts. Even though the
enzyme of A. woodii is highly stable with a temperature optimum of 40 °C,
long-term stability has yet to be determined. Remarkably, the [FeFe] hydrogenase
module of the HDCR is the [FeFe] hydrogenase being completely CO tolerant [54].
In contrast to other [FeFe] hydrogenases where CO leads to irreversible damage to
the H-cluster [55–57], CO inhibition of the [FeFe] hydrogenase module is fully
reversible, even though the enzyme is strongly inhibited by CO [54]. This feature
could be a result of the metabolism of A. woodii, where CO is an intermediate of the
WLP. However, since the HDCR requires strict anoxic conditions and/or a low
redox potential, a biotechnological application is rather difficult. To overcome this
problem, we established a whole-cell system for the efficient hydrogenation of CO 2
to formate [15]. The system is based on the knockout of the ATP-dependent further
conversion of formate in the WLP of acetogens. Formate is transiently excreted and
accumulated during autotrophic growth in A. woodii [58] and accumulation requires
inhibition of the ATP-dependent further conversion of formate in the WLP. By
reducing the cellular energy content, formate is no longer converted to acetate and
Acetogenic Bacteria for Biotechnological Applications
115