Nitrile hydratase

nitrile hydratase
Identifiers
EC number	4.2.1.84
CAS number	82391-37-5
Databases
IntEnz	IntEnz view
BRENDA	BRENDA entry
ExPASy	NiceZyme view
KEGG	KEGG entry
MetaCyc	metabolic pathway
PRIAM	profile
PDB structures	RCSB PDB PDBe PDBsum
Gene Ontology	AmiGO / EGO
Search PMC articles PubMed articles

In enzymology, nitrile hydratases (NHases; EC 4.2.1.84) are mononuclear iron or non-corrinoid cobalt enzymes that catalyse the hydration of diverse nitriles to their corresponding amides

R-C≡N + H₂O → R-C(O)NH₂

Metal cofactor

In biochemistry, cobalt is in general found in a corrin ring, such as in vitamin B₁₂. Nitrile hydratase is one of the rare enzyme types that use cobalt in a non-corrinoid manner. The mechanism by which the cobalt is transported to NHase without causing toxicity is unclear, although a cobalt permease has been identified, which transports cobalt across the cell membrane. The identity of the metal in the active site of a nitrile hydratase can be predicted by analysis of the sequence data of the alpha subunit in the region where the metal is bound. The presence of the amino acid sequence VCTLC indicates a Co-centred NHase and the presence of VCSLC indicates Fe-centred NHase.

Metabolic pathway

Nitrile hydratase and amidase are two hydrating and hydrolytic enzymes responsible for the sequential metabolism of nitriles in bacteria that are capable of utilising nitriles as their sole source of nitrogen and carbon, and in concert act as an alternative to nitrilase activity, which performs nitrile hydrolysis without formation of an intermediate primary amide. A sequence in genome of the eukaryote Monosiga brevicollis that has been suggested to encode for a nitrile hydratase has been identified.^[1]. This is the first hint that nitrile hydratase activity might also occur outside of prokaryotic organisms.

Industrial applications

NHases have been efficiently used for the industrial production of acrylamide from acrylonitrile and for removal of nitriles from wastewater. Photosensitive NHases intrinsically possess nitric oxide (NO) bound to the iron centre, and its photodissociation activates the enzyme.

Structure

Structure of nitrile hydratase.^[2]

NHases are composed of two types of subunits, α and β, which are not related in amino acid sequence. NHases exist as αβ dimers or α₂β₂ tetramers and bind one metal atom per αβ unit. The 3-D structures of a number of NHases have been determined. The α subunit consists of a long extended N-terminal "arm", containing two α-helices, and a C-terminal domain with an unusual four-layered structure (α-β-β-α). The β subunit consists of a long N-terminal loop that wraps around the α subunit, a helical domain that packs with N-terminal domain of the α subunit, and a C-terminal domain consisting of a β-roll and one short helix.

Nitrile hydratase, alpha chain Identifiers Symbol NHase_alpha Pfam PF02979 InterPro IPR004232 SCOP 2ahj Available protein structures: Pfam structures PDB RCSB PDB; PDBe PDBsum structure summary

Nitrile hydratase beta subunit Identifiers Symbol NHase_beta Pfam PF02211 InterPro IPR003168 SCOP 2ahj Available protein structures: Pfam structures PDB RCSB PDB; PDBe PDBsum structure summary

Enzymatic mechanism

The metal centre is located in the central cavity at the interface between two subunits. All protein ligands to the metal atom are provided by the α subunit. The protein ligands to the iron are the sidechains of the three cysteine (Cys) residues and two mainchain amide nitrogens. The metal ion is octahedrally coordinated, with the protein ligands at the five vertices of an octahedron. The sixth position, accessible to the active site cleft, is occupied either by NO or by a solvent-exchangeable ligand (hydroxide or water). The two Cys residues coordinated to the metal are post-translationally modified to Cys-sulfinic (Cys-SO₂H) and -sulfenic (Cys-SOH) acids.

References

1. ^ Foerstner KU, Doerks T, Muller J, Raes J, Bork P (2008). Hannenhalli, Sridhar. ed. "A nitrile hydratase in the eukaryote Monosiga brevicollis". PLoS ONE 3 (12): e3976. doi:10.1371/journal.pone.0003976. PMC 2603476. PMID 19096720. http://www.pubmedcentral.nih.gov/articlerender.fcgi?tool=pmcentrez&artid=2603476. 
2. ^ Nagashima S, Nakasako M, Dohmae N, et al. (May 1998). "Novel non-heme iron center of nitrile hydratase with a claw setting of oxygen atoms". Nat. Struct. Biol. 5 (5): 347–51. doi:10.1038/nsb0598-347. PMID 9586994. 

Further reading

• Prasad, S; Bhalla, TC (May 2010). "Nitrile hydratases (NHases): At the interface of academia and industry .". Biotechnology Advances 28 (6): 725. doi:10.1016/j.biotechadv.2010.05.020. PMID 20685247. 
• Rzeznicka, K; Schätzle, S; Böttcher, D; Klein, J; Bornscheuer, UT (Aug 2009). "Cloning and functional expression of a nitrile hydratase (NHase) from Rhodococcus equi TG328-2 in Escherichia coli, its purification and biochemical characterisation.". Appl Microbiol Biotechnol 85 (5): 1417–25. doi:10.1007/s00253-009-2153-y. PMID 19662400. 
• Song, L; Wang, M; Yang, X; Qian, S (Jun 2007). "Purification and characterization of the enantioselective nitrile hydratase from Rhodococcus sp. AJ270.". Biotechnol J 2 (6): 717–24. doi:10.1002/biot.200600215. PMID 17330219. 
• Miyanaga, A; Fushinobu, S; Ito, K; Shoun, H; Wakagi, T (Jan 2004). "Mutational and structural analysis of cobalt-containing nitrile hydratase on substrate and metal binding.". Eur J Biochem 271 (2): 429–38. doi:10.1046/j.1432-1033.2003.03943.x. PMID 14717710. 
• DiCosimo, EC; Eisenberg, A; Fager, SK; Perkins, NE; Gallagher, FG; Cooper, SM; Gavagan, JE; Stieglitz, B et al. (Oct 1999). "5-Cyanovaleramide production using immobilized Pseudomonas chlororaphis B23.". Bioorg Med Chem 7 (10): 2239–45. doi:10.1016/S0968-0896(99)00157-1. PMID 10579532.