Nitric-oxide reductase

Nitric oxide reductase, an enzyme, catalyzes the reduction of

Nomenclature

Nitric oxide reductase was assigned Enzyme Commission number (EC) 1.7.2.5. Enzyme Commission numbers are the standard naming system used for enzymes.^[5] The EC identifies the class, subclass, sub-subclass, and serial number of the enzyme.^[5] Nitric oxide reductase is in Class 1, therefore it is an oxidoreductases.^[5]

Nitric oxide reductase belongs to the family of oxidoreductases, specifically those acting on other nitrogenous compounds as donors with other acceptors. The systematic name of this enzyme class is nitrous-oxide:acceptor oxidoreductase (NO-forming). Other names in common use include nitrogen oxide reductase, and nitrous-oxide:(acceptor) oxidoreductase (NO-forming).

Function

Organisms reduce nitrate (NO₃⁻) to nitrogen gas (N₂) through the process of denitrification, see Figure 1.^[1][2] Two important intermediates of the reduction pathway are nitric oxide (NO) and nitrous oxide (N₂O).^[1][2] The reducing reaction that transforms NO into N₂O is catalyzed by nitric oxide reductase (NOR).^[1][2][3][4]

NO is reduced to N₂O also to prevent cellular toxicity.^[4][6] N₂O, a potent greenhouse gas, is released.^[1][4]

Reaction

In

2 NO + 2 e⁻ + 2 H⁺ ${\displaystyle \rightleftharpoons }$ N₂O + H₂O[4]

The enzyme acts on 2 nitric oxide (substrate).^[2] The enzyme converts NO, electrons and protons to products: nitrous oxide, and H₂O.^[2]

Inputs: 2 molecules of NO, 2 electrons, 2 protons^[2]

Outputs: 1 molecule of N₂O, 1 molecule of H₂O^[2]

Mechanism

NOR catalyzes the formation of nitrogen to nitrogen (N--N)  bonding.^[1][3][6] The conformation changes of the active site and attached ligands (ie. Glu211) allows NO to be positioned in the crowded binuclear center and form N--N bonds.^[4]

The precise mechanism of catalysis is still unknown, although hypotheses have been proposed.^[3][4]

Cordas et al. 2013 proposes three options: the trans-mechanism, the cis-FeB and the cis-heme b3 mechanisms.^[3]

Based on the structure of the enzyme, Shiro 2012 proposes the following mechanism: (1) NO molecules bind at the binuclear center, (2) electrons are transferred from the ferrous irons to the NO, (3) charged NO molecules have the potential to form N to N bonds, and (4) N to O bonds are potentially broken by water, allowing for the N₂O and H₂O to be released.^[4]

According to Hino et al. 2010, the changing charge of the active site causes NO to bind, form N₂O and leave the enzyme. The NOR active site is positioned near two hydrogen bound glutamic acids (Glu). The Glu groups provide an electron-negative charge to the active site.^[1] The electro-negative charge reduces the reaction potential for heme b3 and allows NO to bind to the binuclear activation site.^[1] Glu residues also provide protons needed for removal of N₂O and production of H₂O.^[1]

Structure

Subunits

NOR is made up of two subunits, NorC (small) and NorB (large), with a binuclear iron centre.^[1][3][4] The binuclear iron center is the active site.^[1][2][3][4] It is composed of two b-type hemes and a non-heme iron (FeB).^[1][2][3][4] The ligands are connected through a μ-oxo bridge.^[3] Histidine (His) residues are attached to the heme b3 in the small subunit.^[1] The hydrophilic region of the larger subunit has His and methionine (Met) ligands.^[1] Structure is similar to cytochrome oxidases.^[1][4]

The active site is conserved between cNOR and qNOR, although differences (ie. heme type) occur between cNOR and qNOR.^[4]

Folding

Enzymatic folding produced 13 alpha-helices (12 from NorB, 1 from NorC) located within and through the membrane.

Bacteria, archaea and fungi use NOR.[4]^[6] qNOR is found in denitrifying bacteria and archaea, as well as pathogenic bacteria not involved in denitrification.^[4] Denitrifying fungi reduce NO using P-450nor soluble enzyme.^[6]

Types

Three types of NOR were identified from bacteria: cNOR, qNOR, and qCuNOR.