PDBsum entry 3hq0

Oxidoreductase

PDB id: 3hq0

Contents

Protein chains

297 a.a. *

Ligands

M3P ×3

Metals

_FE ×4

Waters ×409

* Residue conservation analysis

PDB id:

3hq0

Name:

Oxidoreductase

Title:

Crystal structure analysis of the 2,3-dioxygenase lapb from pseudomonas in complex with a product

Structure:

Catechol 2,3-dioxygenase. Chain: a, b, c, d. Engineered: yes

Source:

Pseudomonas sp. Kl28. Organism_taxid: 237609. Strain: kctc 22206. Expressed in: escherichia coli bl21(de3). Expression_system_taxid: 469008.

Resolution:

2.00Å R-factor: 0.222 R-free: 0.269

Authors:

J.-H.Cho,S.Rhee

Key ref:

J.H.Cho et al. (2009). Crystal structure and functional analysis of the extradiol dioxygenase LapB from a long-chain alkylphenol degradation pathway in Pseudomonas. J Biol Chem, 284, 34321-34330. PubMed id: 19828456 DOI: 10.1074/jbc.M109.031054

Date:

05-Jun-09 Release date: 20-Oct-09

Protein chains

Q7WYF5  (Q7WYF5_9PSED) -  Metapyrocatechase from Pseudomonas alkylphenolica

Seq: 309 a.a.

Struc: 297 a.a.

Enzyme reactions

• Enzyme class: E.C.1.13.11.2 - catechol 2,3-dioxygenase.
• Reaction: catechol + O2 = (2Z,4E)-2-hydroxy-6-oxohexa-2,4-dienoate + H⁺

catechol (Bound ligand (Het Group name = M3P) matches with 72.73% similarity) + O2 = (2Z,4E)-2-hydroxy-6-oxohexa-2,4-dienoate + H(+)

• Cofactor: Fe(2+)

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

Added reference

DOI no: 10.1074/jbc.M109.031054

J Biol Chem 284:34321-34330 (2009)

PubMed id: 19828456

Crystal structure and functional analysis of the extradiol dioxygenase LapB from a long-chain alkylphenol degradation pathway in Pseudomonas.

J.H.Cho, D.K.Jung, K.Lee, S.Rhee.

ABSTRACT

LapB is a non-heme Fe(II)-dependent 2,3-dioxygenase that catalyzes the second step of a long-chain alkylphenol (lap) degradation pathway in Pseudomonas sp. KL28 and belongs to the superfamily of type I extradiol dioxygenases. In this study, the crystal structures of substrate-free LapB and its complexes with a substrate or product were determined, along with a functional analysis of the active site residues. Structural features of the homotetramer are similar to those of other type I extradiol dioxygenases. In particular, the active site is located in the C-domain of each monomer, with a 2-His-1-carboxylate motif as the first coordination shell to iron ion. A comparison of three different structures in the catalytic cycle indicated catalysis-related local conformational changes in the active site. Specifically, the active site loop containing His-248 exhibits positional changes upon binding of the substrate and establishes a hydrogen-bonding network with Tyr-257, which is near the hydroxyl group of the substrate. Kinetic analysis of the mutant enzymes H248A, H248N, and Y257F showed that these three mutant enzymes are inactive, suggesting that this hydrogen-bonding network plays a crucial role in catalysis by deprotonating the incoming substrate and leaving it in a monoanionic state. Additional functional analysis of His-201, by using H201A and H201N mutants, near the dioxygen-binding site also supports its role as base and acid catalyst in the late stage of catalysis. We also noticed a disordered-to-ordered structural transition in the C-terminal region, resulting in the opening or closing of the active site. These results provide detailed insights into the structural and functional features of an extradiol dioxygenase that can accommodate a wide range of alkylcatechols.

Selected figure(s)

Figure 2.
Dimeric and tetrameric interfaces. A, the dimeric interface between subunits A and D are shown, with the interface residues defined using the program CONTACT (17). B, the tetrameric interface was formed mainly by residues belonging to α5.

Figure 3.
Active site of native LapB and its complexes. The active site for substrate-free LapB and its complex with 4MC or product is shown with a stick model, with a schematic representation of the interactions shown in the right panel. The dashed lines indicate putative hydrogen bonds, which are labeled with the average interatomic distance (in Å), and the decorated arcs represent van der Waals interactions of less than 4.0 Å. A, the active site for substrate-free LapB in subunit B is shown. Three water molecules depicted as red spheres occupy the first coordination shell and interact with iron ion, a black sphere. B, a 2F[o] − F[c] electron density map contoured at 1 σ is overlaid on the model of 4MC in the LapB-4MC complex (subunit C). For clarity, Leu-156, which is also a part of the active site, is not shown in this figure and also not shown in C but is displayed in Fig. 1E. C, same as B except that 2-hydroxy-6-oxohepta-2,4-dienoic acid, the product of 3MC produced via proximal cleavage by LapB, is shown with a 2F[o] − F[c] electron density map contoured at 0.8 σ in subunit B.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2009, 284, 34321-34330) copyright 2009.

Figures were selected by an automated process.