nitrogenase

1. STRUCTURE OF NITROGENASE
ENZYME AND ITS EFFICIENCY AT
LOW OXYGEN CONCENTRATION
PRESENTED BY: GROUP 8
JAISHA-E-JANNAT,ALIZA
ASGHAR,AIMAN
IQBAL,MINAHIL
SHAMIM,TAZMEEN NAZIR

2. NITROGENASE ENZYME
 Bacteria can change atmospheric nitrogen into usable
nitrogen in a process called nitrogen fixation.
 An organism that can fix nitrogen is a called a
diazotrope.
 Nitrogen in the atmosphere is pretty much inert
because the 2 nitrogen atoms are joined together in a
triple bond, which takes a lot of energy to break.
 Bacteria have special enzymes called nitrogenase that
will break this bond and convert the nitrogen into
ammonia.

3. NITROGENASE ENZYME
 This enzyme is a highly complex proteins called the
nitrogenase complex; its central components are
 dinitrogenase reductase and
 dinitrogenase
 Dinitrogenase reductase (Mr 60,000) is a dimer of two
identical subunits.
 It contains a single 4Fe-4S redox center, bound between the
subunits.
 It can be oxidized and reduced by one electron. It also has
two binding sites for ATP/ADP (one site on each subunit).

4. NITROGENASE ENZYME
 Dinitrogenase (Mr 240,000), an α2β2 tetramer, has two Fe-
containing cofactors that transfer electrons.
 One, the P cluster, has a pair of 4Fe-4S centers; these share a
sulfur atom, making an 8Fe-7S
 center.
 The second cofactor in dinitrogenase, the FeMo cofactor, is a
novel structure composed of 7 Fe atoms, 9 inorganic S atoms, a
Cys side chain, and a single carbon atom in the center of the FeS
cluster.
 Also part of the cofactor is a molybdenum atom, with ligands
that include three inorganic S atoms, a His side chain, and two
oxygen atoms from a molecule of homocitrate that is an intrinsic
part of the FeMo cofactor.

5. NITROGENASE ENZYME
 There is also a form of nitrogenase that contains
vanadium rather than molybdenum, and some
bacterial species can produce both types.
 The vanadium-containing enzyme may be the
primary nitrogen-fixing system under some
conditions. The vanadium nitrogenase of Azotobacter
vinelandii has theremarkable capacity to catalyze the
reduction of carbon monoxide (CO) to ethylene
(C2H4), ethane, and propane.

6. NITROGENASE ENZYME
 Nitrogen fixation is carried out by a highly reduced form of
dinitrogenase and requires eight electrons: six for the
reduction of N2 and two to produce one molecule of H2.
 Dinitrogenase is reduced by the transfer of electrons from
dinitrogenase reductase. The dinitrogenase tetramer has
two binding sites for the reductase.
 The required eight electrons are transferred from reductase
to dinitrogenase one at a time: a reduced reductase
molecule binds to the dinitrogenase and transfers a single
electron, then the oxidized reductase dissociates from
dinitrogenase, in a repeating cycle.

7. FIGURE : Nitrogenase complex with cofactors. The holoenzyme consists of
- Two identical dinitrogenase reductase molecules (green), each with a 4Fe-4S
redox center and binding sites for two ATP, and Two identical dinitrogenase
heterodimers (purple and blue), each with a P cluster (Fe-S center) and an FeMo
cofactor. In this structure, ADP is bound in the ATP site, to make the crystal
more stable. The FeMo cofactor has a Mo atom with three S ligands, a His
Ligand, and two oxygen ligands from a molecule of homocitrate.

8. ACTIVITY OF NITROGENASE ENZYME AT
LOW OXYGEN CONCENTRATION
 Nitrogenase is extremely sensitive to oxygen. Therefore N2
fixation can proceed only at very low oxygen concentrations. The
nodules form an anaerobic compartment.
 Since N2 fixation depends on the uptake of nitrogen from the
air, the question arises how is the enzyme protected against the
oxygen present in air?
 The answer is that oxygen, which has diffused together with
nitrogen into the nodules, is consumed by the respiratory chain
contained in the bacteroid membrane. Due to a very high
affinity of the bacteroid cytohrome-a/a3 complex, respiration is
still possible with an oxygen concentration of only 10-9 mol/L.

9. ACTIVITY OF NITROGENASE ENZYME AT LOW
OXYGEN CONCENTRATION
 Doubling of the O2 content in air (with a corresponding decrease of the N2 content)
resulted in a doubling of the rate of N2 fixation. But, because of the O2 sensitivity of the
nitrogenase, a further increase in O2 resulted in a steep decline in N2 fixation.
 Since the bacterial respiratory chain is located in the membrane and nitrogenase in the
interior of the bacteroids, O2 is kept at a safe distance from nitrogenase.
 The cells infected by rhizobia form leghemoglobin, which is very similar to the
myoglobin of animals, but has a 10-fold higher affinity for oxygen. The oxygen
concentration required for half saturation of leghemoglobin amounts to only 10–20 ×10-
9 mol/L.
 Leghemoglobin is located in the cytosol of host cell outside the peribacteroid membrane
and present there in unusually high concentrations .
 Leghemoglobin can amount to 25% of the total soluble protein of the nodules and gives
them a pink color.
 It has been proposed that leghemoglobin plays a role in the transport of oxygen within
the nodules. However, it is more likely that it serves as an oxygen buffer to ensure
continuous electron transport in the bacteroids at the very low prevailing O2
concentration in the nodules.