Structure of catechol 1,2-dioxygenase from Pseudomonas arvilla (original) (raw)
Related papers
Acta crystallographica, 2006
Class II extradiol-cleaving catecholic dioxygenase, a key enzyme of aromatic compound degradation in bacteria, cleaves the aromatic ring of catechol by adding two O atoms. PheB is one of the class II extradiol-cleaving catecholic dioxygenases and shows a high substrate specificity for catechol derivatives, which have one aromatic ring. In order to reveal the mechanism of the substrate specificity of PheB, PheB has been crystallized by the hanging-drop vapourdiffusion method using PEG 4000 as a precipitant. The space group of the obtained crystal was P2 1 2 1 2 1 , with unit-cell parameters a = 65.5, b = 119.2, c = 158.7 Å. The crystal diffracted to 2.3 Å resolution.
Homology modeling and docking studies of Catechol-2,3-dioxygenase
Catechol 2, 3-dioxygenase is present in several types of bacteria and undergoes degradation of environmental pollutants through an important key biochemical pathways. Specifically, this enzyme cleaves aromatic rings of several environmental pollutants such as toluene, xylene, naphthalene and biphenyl derivatives. Hence, the importance of Catechol 2, 3-dioxygenase and its role in the degradation of environmental pollutants made us to predict the three-dimensional structure of Catechol 2, 3-dioxygenase from Burkholderia cepacia. The 10ns molecular dynamics simulation was carried out to check the stability of the modeled Catechol 2, 3dioxygenase. The results show that the model was energetically stable, and it attains their equilibrium within 2000 ps of production MD run. The docking of various petroleum hydrocarbons into the Catechol 2,3-dioxygenase reveals that the benzene, Ostrong hydrogen bond and Van der waals interaction with the active site residues of H150, L152, W198, H206, H220, H252, I254, T255, Y261, E271, L276 and F309. Free energy of binding and estimated inhibition constant of these compounds demonstrates that they are energetically stable in their binding cavity. Chrysene shows positive energy of binding in the active site atom of Fe. Except Pyrene all the substrates made close contact with Fe atom by the distance ranges from 1.67 to 2.43 Å. In addition to that, the above mentioned substrate except pyrene all other made π-π stacking interaction with H252 by the distance ranges from 3.40 to 3.90 Å. All these docking results reveal that, except Chrysene all other substrate has good free energy of binding to hold enough in the active site and makes strong VdW interaction with Catechol-2,3-dioxygenase. These results suggest that, the enzyme is capable of catalyzing the above-mentioned substrate.
Journal of Biological …, 2004
Catechol 2,3-dioxygenase (C2,3O) from Pseudomonas stutzeri OX1, which is able to grow on various aromatic substrates as the sole source of carbon and energy, has been expressed in Escherichia coli, purified, characterized, and found to be very similar to other ...
Crystal structure of BphC, a halotolerant catechol dioxygenase
2019
The release of synthetic chemical pollutants in the environment is posing serious health risks. Enzymes, including oxygenases, play a crucial role in xenobiotic degradation. In the present study, we employed a functional metagenomics approach to overcome the limitation of cultivability of microbes under standard laboratory conditions in order to isolate novel dioxygenases capable of degrading recalcitrant pollutants. Fosmid clones possessing dioxygenase activity were further sequenced, and their genes were identified using bioinformatics tools. Two positive fosmid clones, SD3 and RW1, suggested the presence of 2,3-dihydroxybiphenyl 1,2dioxygenase (BphC-SD3) and catechol 2,3-dioxygenase (C23O-RW1), respectively. Recombinant versions of these enzymes were purified to examine their pollutantdegrading abilities. The crystal structure of BphC-SD3 was determined at 2.6-Å resolution, revealing a two-domain architecture, i.e., N-terminal and C-terminal domains, with the sequential arrangement of ␣ in each domain, characteristic of Fe-dependent class II type I extradiol dioxygenases. The structure also reveals the presence of conserved amino acids lining the catalytic pocket and Fe 3ϩ metal ion in the large funnel-shaped active site in the C-terminal domain. Further studies suggest that Fe 3ϩ bound in the BphC-SD3 active site probably imparts aerobic stability. We further demonstrate the potential application of BphC-SD3 in biosensing of catecholic compounds. The halotolerant and oxygen-resistant properties of these enzymes reported in this study make them potential candidates for bioremediation and biosensing applications. IMPORTANCE The disposal and degradation of xenobiotic compounds have been serious issues due to their recalcitrant properties. Microbial oxygenases are the fundamental enzymes involved in biodegradation that oxidize the substrate by transferring oxygen from molecular oxygen. Among oxygenases, catechol dioxygenases are more versatile in biodegradation and are well studied among the bacterial world. The use of catechol dioxygenases in the field is currently not practical due to their aerobically unstable nature. The significance of our research lies in the discovery of aerobically stable and halotolerant catechol dioxygenases that are efficient in degrading the targeted environmental pollutants and, hence, could be used as costeffective alternatives for the treatment of hypersaline industrial effluents. Moreover, the structural determination of novel catechol dioxygenases would greatly enhance our knowledge of the function of these enzymes and facilitate directed evolution to further enhance or engineer desired properties.
Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics, 2011
Intradiol-cleaving catechol 1,2 dioxygenases are Fe(III) dependent enzymes that act on catechol and substituted catechols, including chlorocatechols pollutants, by inserting molecular oxygen in the aromatic ring. Members of this class are the object of intense biochemical investigations aimed at the understanding of their catalytic mechanism, particularly for designing mutants with selected catalytic properties. We report here an in depth investigation of catechol 1,2 dioxygenase IsoB from Acinetobacter radioresistens LMG S13 and its A72G and L69A mutants. By applying a multidisciplinary approach that includes high resolution X-rays crystallography, mass spectrometry and single crystal microspectrophotometry, we characterised the phospholipid bound to the enzyme and provided a structural framework to understand the inversion of substrate specificity showed by the mutants. Our results might be of help for the rational design of enzyme mutants showing a biotechnologically relevant substrate specificity, particularly to be used in bioremediation. This article is part of a Special Issue entitled: Protein Structure and Function in the Crystalline State.
Experimental Results
This manuscript reports structure–function studies of Catechol 2,3-dioxygenase (C23O64), which is the second enzyme in the metabolic degradation pathway of 3-nitrotoluene by Diaphorobacter sp. strain DS2. The recombinant protein is a ring cleavage enzyme for 3-methylcatechol and 4-methylcatechol products formed after dioxygenation of the aromatic ring. Here we report the substrate-free, substrate-bound, and substrate-analog bound crystal structures of C23O64. The protein crystallizes in the P6(2)22 space-group. The structures were determined by molecular replacement and refined to resolutions of 2.4, 2.4, 2.2 Å, respectively. A comparison of the structures with related extradiol dioxygenases showed 22 conserved residues. A comparison of the active site pocket with catechol 2,3-dioxygenase (LapB) from Pseudomonas sp KL28 and homoprotocatechuate 2,3-dioxygenase (HPCD) from Brevibacterium fuscum shows significant similarities to suggest that the mechanism of enzyme action is similar to...
Intersubunit interaction and catalytic activity of catechol 2,3-dioxygenases
Biochemical Journal, 2003
Catechol 2,3-dioxygenases (C23Os; EC 1.3.11.2) form a large protein family that is divided into several subgroups. Amino acid sequences of C23Os belonging to subgroup I.2.A and those belonging to I.2.B are found to be approx. 50% identical. When the central parts of the C23O sequences belonging to I.2.B were fused with the N-terminal and C-terminal sequences of I.2.A C23O, the hybrid enzymes were not active. To understand why these hybrid C23Os were inactive, hybrids between XylEP (C23O found in a Pseudomonas strain; subgroup I.2.A) and XylES (C23O found in a Sphingomonas strain; subgroup I.2.B) were constructed. HB3-C23O consisted mostly of the XylES sequence, except that its C-terminal end was derived from XylEP. While HB3-C23O was not active, HB4-C23O, carrying shorter C-terminal XylEP sequences than HB3-C23O, was active. This observation indicated that certain amino acid residues at the C-terminus were crucial for C23O activity in the hybrid forms of enzymes between XylEP and Xy...
Activity of a carboxyl-terminal truncated form of catechol 2,3-dioxygenase from Planococcus sp. S5
TheScientificWorldJournal, 2014
Catechol 2,3-dioxygenases (C23Os, E.C.1.13.12.2) are two domain enzymes that catalyze degradation of monoaromatic hydrocarbons. The catalytically active C-domain of all known C23Os comprises ferrous ion ligands as well as residues forming active site pocket. The aim of this work was to examine and discuss the effect of nonsense mutation at position 289 on the activity of catechol 2,3-dioxygenase from Planococcus strain. Although the mutant C23O showed the same optimal temperature for activity as the wild-type protein (35 °C), it exhibited activity slightly more tolerant to alkaline pH. Mutant enzyme exhibited also higher affinity to catechol as a substrate. Its K(m) (66.17 µM) was approximately 30% lower than that of wild-type enzyme. Interestingly, removal of the C-terminal residues resulted in 1.5- to 1.8-fold (P < 0.05) increase in the activity of C23OB61 against 4-methylcatechol and 4-chlorocatechol, respectively, while towards catechol the activity of the protein dropped to ...
FEBS Letters, 1994
The interaction of different classes of inhibitors with the extradiol cleaving catechol2,3-dioxygenase from Pseudomonasputida mt-2 was monitored by longitudinal and transverse proton relaxation measurements as well as by kinetic activity studies in order to characterize the type of interaction of such inhibitors with the active site of the enzyme. The average distances of the inhibitors from the catalytic iron(I1) ion have been estimated from the 'H longitudinal relaxation rates. Phenols and aliphatic ketones appear to be coordinated to the iron(I1) ion in the active site.