[FeFe]-hydrogenases are often involved in the reduction of protons to produce
H 2 . These are the only type of hydrogenases found in the eukaryotic microorganisms (Vignais and Colbeau 2004). In green microalgae they are located exclusively
in the stroma of the chloroplast (Eroglu and Melis 2011). These hydrogenases are
monomeric or dimeric with an average molecular weight of 50 kDa. The active site
cluster of the enzyme also known as H-cluster consists of six Fe atoms arranged as
[4Fe-4S] sub-cluster to which [2Fe-2S] extension is covalently bridged via cysteine
residue. The Fe atoms of the active site are bound to non-protein ligands, CN
− and
CO groups (Peters et al. 1998, 2015). The H-cluster of the [FeFe]-hydrogenases
makes them different from the other H 2 -producing enzymes and results in 100-fold
higher enzyme activity (Happe et al. 2002). However, in spite of high specific
activity these enzymes get easily inactivated by O 2 or CO 2 . The green microalgae,
C. reinhardtii, encodes two [FeFe]-hydrogenases (HydA1 and HydA2) which are
74% similar and are expressed under anaerobic condition (Forestier et al. 2003).
[NiFe]-hydrogenases are the most numerous hydrogenases found only in
prokaryotes: cyanobacteria, bacteria and archaea. The core enzyme consists of the
a–b heterodimer, where the larger a-subunit possesses the NiFe bimetallic centre
and the smaller b-subunit consists of the Fe–S clusters which transfer the electrons
from the active site to the e
− acceptor molecule (Kim and Kim 2011). In the active
site, presence of non-protein ligands (CN
− and CO groups) bound to the Fe atom is
the common structural characteristic of the [FeFe]- and [NiFe]-hydrogenases
(Peters et al. 2015). In cyanobacterial species, these enzymes occur in two different
types: hup-encoded [NiFe]-uptake hydrogenases and hox-encoded [NiFe]-bidirectional hydrogenases. Uptake hydrogenase catalyses the oxidation of H 2 to recover
the energy lost during N 2 fixation. These are found in all nitrogen-fixing
cyanobacteria, but their presence in non-nitrogen-fixing cyanobacteria is still
under question (Tamagnini et al. 2002). The small subunit of the enzyme does not
contain the signal peptide at N-terminal; therefore, the enzyme is localized on the
cytoplasmic side of either the cytoplasmic or thylakoid membrane (Tamagnini et al.
2002). In the filamentous cyanobacteria, these enzymes are found in the thylakoid
membrane of the heterocysts (Tiwari and Pandey 2012). Inactivation of the gene
(hupS) encoding the small subunit of uptake hydrogenase led to the enhanced and
sustained H 2 production in Anabaena siamensis TISTR 8012 under high light
intensity (Khetkorn et al. 2012). Bidirectional hydrogenase is the reversible enzyme
that can either evolve or consume H 2 according to the existing redox state of the
cell’s photosynthetic membrane (Eroglu and Melis 2011). This enzyme is present in
both nitrogen-fixing and non-nitrogen-fixing cyanobacteria. The enzyme is multimeric because the dimeric module of the enzyme is associated with other subunits
that can bind cofactors. In cyanobacteria, during the period of adaptation to higher
light intensities the reversible hydrogenases may act as an electron valve (Vignais
and Colbeau 2004).
10 Biofuels from Microalgae: Biohydrogen
209
H 2 . These are the only type of hydrogenases found in the eukaryotic microorganisms (Vignais and Colbeau 2004). In green microalgae they are located exclusively
in the stroma of the chloroplast (Eroglu and Melis 2011). These hydrogenases are
monomeric or dimeric with an average molecular weight of 50 kDa. The active site
cluster of the enzyme also known as H-cluster consists of six Fe atoms arranged as
[4Fe-4S] sub-cluster to which [2Fe-2S] extension is covalently bridged via cysteine
residue. The Fe atoms of the active site are bound to non-protein ligands, CN
− and
CO groups (Peters et al. 1998, 2015). The H-cluster of the [FeFe]-hydrogenases
makes them different from the other H 2 -producing enzymes and results in 100-fold
higher enzyme activity (Happe et al. 2002). However, in spite of high specific
activity these enzymes get easily inactivated by O 2 or CO 2 . The green microalgae,
C. reinhardtii, encodes two [FeFe]-hydrogenases (HydA1 and HydA2) which are
74% similar and are expressed under anaerobic condition (Forestier et al. 2003).
[NiFe]-hydrogenases are the most numerous hydrogenases found only in
prokaryotes: cyanobacteria, bacteria and archaea. The core enzyme consists of the
a–b heterodimer, where the larger a-subunit possesses the NiFe bimetallic centre
and the smaller b-subunit consists of the Fe–S clusters which transfer the electrons
from the active site to the e
− acceptor molecule (Kim and Kim 2011). In the active
site, presence of non-protein ligands (CN
− and CO groups) bound to the Fe atom is
the common structural characteristic of the [FeFe]- and [NiFe]-hydrogenases
(Peters et al. 2015). In cyanobacterial species, these enzymes occur in two different
types: hup-encoded [NiFe]-uptake hydrogenases and hox-encoded [NiFe]-bidirectional hydrogenases. Uptake hydrogenase catalyses the oxidation of H 2 to recover
the energy lost during N 2 fixation. These are found in all nitrogen-fixing
cyanobacteria, but their presence in non-nitrogen-fixing cyanobacteria is still
under question (Tamagnini et al. 2002). The small subunit of the enzyme does not
contain the signal peptide at N-terminal; therefore, the enzyme is localized on the
cytoplasmic side of either the cytoplasmic or thylakoid membrane (Tamagnini et al.
2002). In the filamentous cyanobacteria, these enzymes are found in the thylakoid
membrane of the heterocysts (Tiwari and Pandey 2012). Inactivation of the gene
(hupS) encoding the small subunit of uptake hydrogenase led to the enhanced and
sustained H 2 production in Anabaena siamensis TISTR 8012 under high light
intensity (Khetkorn et al. 2012). Bidirectional hydrogenase is the reversible enzyme
that can either evolve or consume H 2 according to the existing redox state of the
cell’s photosynthetic membrane (Eroglu and Melis 2011). This enzyme is present in
both nitrogen-fixing and non-nitrogen-fixing cyanobacteria. The enzyme is multimeric because the dimeric module of the enzyme is associated with other subunits
that can bind cofactors. In cyanobacteria, during the period of adaptation to higher
light intensities the reversible hydrogenases may act as an electron valve (Vignais
and Colbeau 2004).
10 Biofuels from Microalgae: Biohydrogen
209