In enzymology, an azobenzene reductase also known as azoreductase is an enzyme

Enzyme

Enzymes are proteins that catalyze chemical reactions. In enzymatic reactions, the molecules at the beginning of the process, called substrates, are converted into different molecules, called products. Almost all chemical reactions in a biological cell need enzymes in order to occur at rates...

 that catalyzes

Catalysis

Catalysis is the change in rate of a chemical reaction due to the participation of a substance called a catalyst. Unlike other reagents that participate in the chemical reaction, a catalyst is not consumed by the reaction itself. A catalyst may participate in multiple chemical transformations....

 the chemical reaction

Chemical reaction

A chemical reaction is a process that leads to the transformation of one set of chemical substances to another. Chemical reactions can be either spontaneous, requiring no input of energy, or non-spontaneous, typically following the input of some type of energy, such as heat, light or electricity...

:

N,N-dimethyl-1,4-phenylenediamine + aniline + NADP⁺ 4-(dimethylamino)azobenzene + NADPH + H⁺

The 3 substrates

Substrate (biochemistry)

In biochemistry, a substrate is a molecule upon which an enzyme acts. Enzymes catalyze chemical reactions involving the substrate. In the case of a single substrate, the substrate binds with the enzyme active site, and an enzyme-substrate complex is formed. The substrate is transformed into one or...

 of this enzyme are N,N-dimethyl-1,4-phenylenediamine

P-Phenylenediamine

p-Phenylenediamine is an organic compound with the formula C6H42. This derivative of aniline is a colorless solid, but typically samples can contain yellowish impurities arising from oxidation. It is mainly used as a component of engineering polymers and composites. It is also an ingredient in...

, aniline

Aniline

Aniline, phenylamine or aminobenzene is an organic compound with the formula C6H5NH2. Consisting of a phenyl group attached to an amino group, aniline is the prototypical aromatic amine. Being a precursor to many industrial chemicals, its main use is in the manufacture of precursors to polyurethane...

, and NADP⁺

Nicotinamide adenine dinucleotide phosphate

Nicotinamide adenine dinucleotide phosphate, abbreviated NADP or TPN in older notation , is a coenzyme used in anabolic reactions, such as lipid and nucleic acid synthesis, which require NADPH as a reducing agent....

, whereas its 3 products

Product (chemistry)

Product are formed during chemical reactions as reagents are consumed. Products have lower energy than the reagents and are produced during the reaction according to the second law of thermodynamics. The released energy comes from changes in chemical bonds between atoms in reagent molecules and...

 are 4-(dimethylamino)azobenzene

Azobenzene

Azobenzene is a chemical compound composed of two phenyl rings linked by a N=N double bond. It is the best known example of an azo compound. The term 'azobenzene' or simply 'azo' is often used to refer to a wide class of molecules that share the core azobenzene structure, with different chemical...

, NADPH

Nicotinamide adenine dinucleotide phosphate

Nicotinamide adenine dinucleotide phosphate, abbreviated NADP or TPN in older notation , is a coenzyme used in anabolic reactions, such as lipid and nucleic acid synthesis, which require NADPH as a reducing agent....

, and H⁺

Hydrogen ion

Hydrogen ion is recommended by IUPAC as a general term for all ions of hydrogen and its isotopes.Depending on the charge of the ion, two different classes can be distinguished: positively charged ions and negatively charged ions....

.

This enzyme belongs to the family of oxidoreductase

Oxidoreductase

In biochemistry, an oxidoreductase is an enzyme that catalyzes the transfer of electrons from one molecule to another...

s, specifically those acting on other nitrogenous compounds as donors with NAD+ or NADP+ as acceptor.

Mechanism

The reaction catalyzed by this enzyme proceeds via a ping-pong mechanism by using 2 equivalents of NAD(P)H to reduce one equivalent of the azo compound

Azo compound

Azo compounds are compounds bearing the functional group R-N=N-R', in which R and R' can be either aryl or alkyl. IUPAC defines azo compounds as: "Derivatives of diazene , HN=NH, wherein both hydrogens are substituted by hydrocarbyl groups, e.g. PhN=NPh azobenzene or diphenyldiazene." The more...

 substrate (for example methyl red

Methyl red

Methyl red, also called C.I. Acid Red 2, is an indicator dye that turns red in acidic solutions. It is an azo dye, and is a dark red crystalline powder....

 where Ar = p-dimethylaniline

Dimethylaniline

N,N-Dimethylaniline is an organic chemical compound, a substituted derivative of aniline. It consists of a tertiary amine, featuring dimethylamino group attached to a phenyl group. This oily liquid is colourless when pure, but commercial samples are often yellow...

 and Ar' = o-benzoic acid

Benzoic acid

Benzoic acid , C7H6O2 , is a colorless crystalline solid and the simplest aromatic carboxylic acid. The name derived from gum benzoin, which was for a long time the only source for benzoic acid. Its salts are used as a food preservative and benzoic acid is an important precursor for the synthesis...

) into two equivalents of aniline

Aniline

Aniline, phenylamine or aminobenzene is an organic compound with the formula C6H5NH2. Consisting of a phenyl group attached to an amino group, aniline is the prototypical aromatic amine. Being a precursor to many industrial chemicals, its main use is in the manufacture of precursors to polyurethane...

 product:

Ar–N=N–Ar' + 2(NAD(P)H + H⁺) Ar–NH₂ + NH₂–Ar' + 2NAD(P)⁺

Substrate specificity

Most azoreductase isoenzymes can reduce methyl red, but are not able to reduce sulfonated

Aromatic sulfonation

Aromatic sulfonation is an organic reaction in which a hydrogen atom on an arene is replaced by a sulfonic acid functional group in an electrophilic aromatic substitution.-Mechanism:...

 azo dyes. The unique azoreductase isozyme

Isozyme

Isozymes are enzymes that differ in amino acid sequence but catalyze the same chemical reaction. These enzymes usually display different kinetic parameters Isozymes (also known as isoenzymes) are enzymes that differ in amino acid sequence but catalyze the same chemical reaction. These enzymes...

 from Bacillus

Bacillus

Bacillus is a genus of Gram-positive, rod-shaped bacteria and a member of the division Firmicutes. Bacillus species can be obligate aerobes or facultative anaerobes, and test positive for the enzyme catalase. Ubiquitous in nature, Bacillus includes both free-living and pathogenic species...

sp. B29 has the ability to reduce sulfonated azo dyes however.

Nomenclature

The systematic name of this enzyme class is N,N-dimethyl-1,4-phenylenediamine, aniline:NADP+ oxidoreductase. Other names in common use include:

• azo reductase,
• azoreductase,
• azo-dye reductase,
• dibromopropylaminophenylazobenzoic azoreductase,
• dimethylaminobenzene reductase,
• methyl red azoreductase,
• N,N-dimethyl-4-phenylazoaniline azoreductase,
• NAD(P)H:1-(4'-sulfophenylazo)-2-naphthol oxidoreductase,
• NADPH2-dependent azoreductase,
• NADPH2:4-(dimethylamino)azobenzene oxidoreductase,
• NC-reductase,
• new coccine (NC)-reductase,
• nicotinamide adenine dinucleotide (phosphate) azoreductase,
• orange I azoreductase,
• orange II azoreductase,
• p-aminoazobenzene reductase, and
• p-dimethylaminoazobenzene azoreductase.

Structural studies

As of late 2007, 3 structures

Tertiary structure

In biochemistry and molecular biology, the tertiary structure of a protein or any other macromolecule is its three-dimensional structure, as defined by the atomic coordinates.-Relationship to primary structure:...

 have been solved for this class of enzymes, with PDB

Protein Data Bank

The Protein Data Bank is a repository for the 3-D structural data of large biological molecules, such as proteins and nucleic acids....

accession codes , , and . Please check the last updated data on RCSB PDB site.