PDBsum entry 1iun

Hydrolase

PDB id: 1iun

Contents

Protein chains

272 a.a. *

Ligands

ACT

Waters ×115

* Residue conservation analysis

PDB id:

1iun

Name:

Hydrolase

Title:

Meta-cleavage product hydrolase from pseudomonas fluorescens ip01 (cumd) s103a mutant hexagonal

Structure:

Meta-cleavage product hydrolase. Chain: a, b. Synonym: 2-hydroxy-6-oxo-7-methylocta-2,4-dienoate hydrolase. Engineered: yes. Mutation: yes

Source:

Pseudomonas fluorescens. Organism_taxid: 294. Strain: ip01. Gene: cumd. Expressed in: escherichia coli. Expression_system_taxid: 562.

Resolution:

2.80Å R-factor: 0.196 R-free: 0.251

Authors:

S.Fushinobu,T.Saku,M.Hidaka,S.-Y.Jun,H.Nojiri,H.Yamane,H.Shoun, T.Omori,T.Wakagi

Key ref:

S.Fushinobu et al. (2002). Crystal structures of a meta-cleavage product hydrolase from Pseudomonas fluorescens IP01 (CumD) complexed with cleavage products. Protein Sci, 11, 2184-2195. PubMed id: 12192074 DOI: 10.1110/ps.0209602

Date:

06-Mar-02 Release date: 18-Sep-02

Protein chains

P96965  (P96965_PSEFL) -  2-hydroxy-6-oxo-7-methylocta-2,4-dienoate hydrolase from Pseudomonas fluorescens

Seq: 282 a.a.

Struc: 272 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

Enzyme reactions

• Enzyme class: E.C.3.7.1.9 - 2-hydroxymuconate-6-semialdehyde hydrolase.
• Reaction: (2Z,4E)-2-hydroxy-6-oxohexa-2,4-dienoate + H2O = 2-oxopent-4-enoate + formate + H⁺

(2Z,4E)-2-hydroxy-6-oxohexa-2,4-dienoate + H2O = 2-oxopent-4-enoate + formate (Bound ligand (Het Group name = ACT) matches with 75.00% similarity) + H(+)

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

Added reference

DOI no: 10.1110/ps.0209602

Protein Sci 11:2184-2195 (2002)

PubMed id: 12192074

Crystal structures of a meta-cleavage product hydrolase from Pseudomonas fluorescens IP01 (CumD) complexed with cleavage products.

S.Fushinobu, T.Saku, M.Hidaka, S.Y.Jun, H.Nojiri, H.Yamane, H.Shoun, T.Omori, T.Wakagi.

ABSTRACT

2-Hydroxy-6-oxo-7-methylocta-2,4-dienoate hydrolase (CumD) from Pseudomonas fluorescens IP01 hydrolyzes a meta-cleavage product generated in the cumene (isopropylbenzene) degradation pathway. The crystal structures of the inactive S103A mutant of the CumD enzyme complexed with isobutyrate and acetate ions were determined at 1.6 and 2.0 A resolution, respectively. The isobutyrate and acetate ions were located at the same position in the active site, and occupied the site for a part of the hydrolysis product with CumD, which has the key determinant group for the substrate specificity of related hydrolases. One of the oxygen atoms of the carboxyl group of the isobutyrate ion was hydrogen bonded with a water molecule and His252. Another oxygen atom of the carboxyl group was situated in an oxyanion hole formed by the two main-chain N atoms. The isopropyl group of the isobutyric acid was recognized by the side-chains of the hydrophobic residues. The substrate-binding pocket of CumD was long, and the inhibition constants of various organic acids corresponded well to it. In comparison with the structure of BphD from Rhodococcus sp. RHA1, the structural basis for the substrate specificity of related hydrolases, is revealed.

Selected figure(s)

Figure 1.
Fig. 1. Degradation pathway for cumene (A), toluene (B), ethylbenzene (C), and biphenyl (D). Chemical designations are as follows: compound I, cumene; compound II, 3-isopropylcatechol; compound III, 2-hydroxy-6-oxo-7-methylocta-2,4-dienoate (6-isopropyl-HODA); compound IV, 2-hydroxypenta-2,4-dienoate; compound V, toluene; compound VI, 2-hydroxy-6-oxohepta-2,4-dienoate (6-methyl-HODA); compound VII, ethylbenzene; compound VIII, 2-hydroxy-6-oxoocta-2,4-dienoate (6-ethyl-HODA); compound IX, biphenyl; and compound X, 2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoate (6-phenyl-HODA). Enzyme designations are as follows: A, aromatic ring dioxygenase; B, dihydrodiol dehydrogenase; C, extradiol dioxygenase; and D, meta-cleavage compound hydrolase (HODA hydrolase). Compounds III, VI, and VIII serve as good substrates for CumD, whereas compound X does not.

Figure 4.
Fig. 4. (A) Schematic drawing of the atoms and interactions involved in the recognition of ISB300. (B) Superimpositioning of the sigma-A weighted 2F[o]-F[c] electron density maps contoured at 2 and the final model structures of type-II ACT and type-II ISB at the bound acetate and isobutyrate ions. The maps of the type-II ACT and type-II ISB structures are shown in red and blue, respectively. The carbon atoms in the model structures of type-II ACT and type-II ISB are shown in orange and yellow, respectively. Water molecules are represented as green spheres. (C, D) Hydrophobic residues involved in the recognition of the isopropyl group of ISB300. A-weighted 2F[o]-F[c] electron density maps contoured at 2 are shown. The view in D is rotated 90° around the perpendicular axis from that in C.

The above figures are reprinted by permission from the Protein Society: Protein Sci (2002, 11, 2184-2195) copyright 2002.

Figures were selected by the author.