PDBsum entry 2zpi

Lyase

PDB id: 2zpi

Contents

Protein chains

191 a.a. *

211 a.a. *

Ligands

TB0

TRS

Metals

_MG ×4

_FE

Waters ×546

* Residue conservation analysis

PDB id:

2zpi

Name:

Lyase

Title:

Complex of fe-type nitrile hydratase with tert-butylisonitrile, photo- activated for 440min at 293k

Structure:

Nitrile hydratase subunit alpha. Chain: a. Synonym: nitrilase, nhase. Nitrile hydratase subunit beta. Chain: b. Synonym: nitrilase, nhase. Ec: 4.2.1.84

Source:

Rhodococcus erythropolis. Organism_taxid: 1833. Strain: n771. Strain: n771

Resolution:

1.49Å R-factor: 0.158 R-free: 0.182

Authors:

K.Hashimoto,H.Suzuki,K.Taniguchi,T.Noguchi,M.Yohda,M.Odaka

Key ref:

K.Hashimoto et al. (2008). Catalytic Mechanism of Nitrile Hydratase Proposed by Time-resolved X-ray Crystallography Using a Novel Substrate, tert-Butylisonitrile. J Biol Chem, 283, 36617-36623. PubMed id: 18948265 DOI: 10.1074/jbc.M806577200

Date:

11-Jul-08 Release date: 21-Oct-08

Protein chain

P13448  (NHAA_RHOER) -  Nitrile hydratase subunit alpha from Rhodococcus erythropolis

Seq: 207 a.a.

Struc: 191 a.a.*

Protein chain

P13449  (NHAB_RHOER) -  Nitrile hydratase subunit beta from Rhodococcus erythropolis

Seq: 212 a.a.

Struc: 211 a.a.

* PDB and UniProt seqs differ at 2 residue positions (black crosses)

Enzyme reactions

• Enzyme class: Chains A, B: E.C.4.2.1.84 - nitrile hydratase.
• Reaction: an aliphatic primary amide = an aliphatic nitrile + H2O

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

Key reference

DOI no: 10.1074/jbc.M806577200

J Biol Chem 283:36617-36623 (2008)

PubMed id: 18948265

Catalytic Mechanism of Nitrile Hydratase Proposed by Time-resolved X-ray Crystallography Using a Novel Substrate, tert-Butylisonitrile.

K.Hashimoto, H.Suzuki, K.Taniguchi, T.Noguchi, M.Yohda, M.Odaka.

ABSTRACT

Nitrile hydratases (NHases) have an unusual iron or cobalt catalytic center with two oxidized cysteine ligands, cysteine-sulfinic acid and cysteine-sulfenic acid, catalyzing the hydration of nitriles to amides. Recently, we found that the NHase of Rhodococcus erythropolis N771 exhibited an additional catalytic activity, converting tert-butylisonitrile (tBuNC) to tert-butylamine. Taking advantage of the slow reactivity of tBuNC and the photoreactivity of nitrosylated NHase, we present the first structural evidence for the catalytic mechanism of NHase with time-resolved x-ray crystallography. By monitoring the reaction with attenuated total reflectance-Fourier transform infrared spectroscopy, the product from the isonitrile carbon was identified as a CO molecule. Crystals of nitrosylated inactive NHase were soaked with tBuNC. The catalytic reaction was initiated by photo-induced denitrosylation and stopped by flash cooling. tBuNC was first trapped at the hydrophobic pocket above the iron center and then coordinated to the iron ion at 120 min. At 440 min, the electron density of tBuNC was significantly altered, and a new electron density was observed near the isonitrile carbon as well as the sulfenate oxygen of alphaCys(114). These results demonstrate that the substrate was coordinated to the iron and then attacked by a solvent molecule activated by alphaCys(114)-SOH.

Selected figure(s)

Figure 5.
The steric hindrance at Sγ of βMet40 caused by tBuNC. The refined structure around βMet^40 in the nitrosylated NHase without (A) and with (B) tBuNC. Yellow, blue, red, and green spheres represent carbon, nitrogen, oxygen, and sulfur atoms, respectively. The black and red dashed lines indicate the distances between Sγ of βMet^40 and the isonitrile carbon and between Sγ of βMet^40 and the amide oxygen of βMet^40.

Figure 6.
Proposed catalytic mechanisms of NHase. A, isonitrile hydrolysis. B, nitrile hydration.

The above figures are reprinted from an Open Access publication published by the ASBMB: J Biol Chem (2008, 283, 36617-36623) copyright 2008.

Figures were selected by an automated process.