Nitrogenases
Nitrogenase is present in cyanobacteria which catalyses the nitrogen fixation by
reducing the molecular nitrogen into ammonium ions that can be easily utilized by
the organisms. Nitrogen fixation is ATP-requiring irreversible reaction and is
essential for the maintenance of the nitrogen cycle in the atmosphere. The reduction
of nitrogen to ammonia by nitrogenase is accompanied by the reduction of protons
(H
+ ) leading to H 2 production. Nitrogenases are the metalloenzyme, and depending
upon the type of metal cofactor present at the catalytic site, they can be of three
types: molybdenum, iron or vanadium nitrogenases. All these three variants of
nitrogenases are capable to carry out the H 2 production during the nitrogen fixation
but with variable stoichiometries (Eqs. 5–7). However, in the absence of nitrogen,
nitrogenases can exclusively produce the H 2 as described in Eq. 4.
Mo-nitrogenase : N 2 þ 8e
À
þ 8H
þ
! 2NH 3 þ H 2
ð5Þ
Fe-nitrogenase : N 2 þ 21e
À
þ 21H
þ
! 2NH 3 þ 7:5H 2
ð6Þ
Fe-nitrogenase : N 2 þ 12e
À
þ 12H
þ
! 2NH 3 þ 3H 2
ð7Þ
Among all the nitrogenases, the most studied one is molybdenum nitrogenase. It
consists of two proteins: the larger dinitrogenase (Mo–Fe–S protein or protein I)
and the smaller dinitrogenase reductase (Fe–S protein or protein II). The dinitrogenase complex has an average molecular weight of 230 kDa and is a
a 2 b 2 heterotetramer encoded by the nifK and nifD genes. The dinitrogenase
reductase subunit is a homodimer of around 65 kDa and is encoded by nifH gene.
Reductase protein receives the electron either from flavodoxin or ferredoxin (external e
− donor) and transfers it to dinitrogenase protein with concomitant
hydrolysis of ATP. Hydrogen produced by the nitrogenase activity is generally
consumed by the uptake hydrogenases due to which the net H 2 evolution by
cyanobacteria is barely observed, at least in aerobic condition (Almon and Bӧger
1988).
2.2 Dark Fermentation Using Microalgal Biomass
as Feedstock
2.2.1 Anaerobic Fermentation Process
Dark fermentation for bioH 2 production is considered as a promising technology
mainly due to following reasons: process simplicity, no requirement of light energy,
higher rate of H 2 evolution and potentiality to utilize wide variety of substrates
(different biomass and wastewater) for H 2 production. This process involves the
anaerobic breakdown of the high molecular weight organic substrates
210
H. Singh and D. Das
Nitrogenase is present in cyanobacteria which catalyses the nitrogen fixation by
reducing the molecular nitrogen into ammonium ions that can be easily utilized by
the organisms. Nitrogen fixation is ATP-requiring irreversible reaction and is
essential for the maintenance of the nitrogen cycle in the atmosphere. The reduction
of nitrogen to ammonia by nitrogenase is accompanied by the reduction of protons
(H
+ ) leading to H 2 production. Nitrogenases are the metalloenzyme, and depending
upon the type of metal cofactor present at the catalytic site, they can be of three
types: molybdenum, iron or vanadium nitrogenases. All these three variants of
nitrogenases are capable to carry out the H 2 production during the nitrogen fixation
but with variable stoichiometries (Eqs. 5–7). However, in the absence of nitrogen,
nitrogenases can exclusively produce the H 2 as described in Eq. 4.
Mo-nitrogenase : N 2 þ 8e
À
þ 8H
þ
! 2NH 3 þ H 2
ð5Þ
Fe-nitrogenase : N 2 þ 21e
À
þ 21H
þ
! 2NH 3 þ 7:5H 2
ð6Þ
Fe-nitrogenase : N 2 þ 12e
À
þ 12H
þ
! 2NH 3 þ 3H 2
ð7Þ
Among all the nitrogenases, the most studied one is molybdenum nitrogenase. It
consists of two proteins: the larger dinitrogenase (Mo–Fe–S protein or protein I)
and the smaller dinitrogenase reductase (Fe–S protein or protein II). The dinitrogenase complex has an average molecular weight of 230 kDa and is a
a 2 b 2 heterotetramer encoded by the nifK and nifD genes. The dinitrogenase
reductase subunit is a homodimer of around 65 kDa and is encoded by nifH gene.
Reductase protein receives the electron either from flavodoxin or ferredoxin (external e
− donor) and transfers it to dinitrogenase protein with concomitant
hydrolysis of ATP. Hydrogen produced by the nitrogenase activity is generally
consumed by the uptake hydrogenases due to which the net H 2 evolution by
cyanobacteria is barely observed, at least in aerobic condition (Almon and Bӧger
1988).
2.2 Dark Fermentation Using Microalgal Biomass
as Feedstock
2.2.1 Anaerobic Fermentation Process
Dark fermentation for bioH 2 production is considered as a promising technology
mainly due to following reasons: process simplicity, no requirement of light energy,
higher rate of H 2 evolution and potentiality to utilize wide variety of substrates
(different biomass and wastewater) for H 2 production. This process involves the
anaerobic breakdown of the high molecular weight organic substrates
210
H. Singh and D. Das