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Urszula Guzik - One of the best experts on this subject based on the ideXlab platform.
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characterization of Catechol 2 3 dioxygenase from planococcus sp strain s5 induced by high phenol concentration
Acta Biochimica Polonica, 2012Co-Authors: Katarzyna Hupertkocurek, Urszula Guzik, Danuta WojcieszynskaAbstract:This study aimed at characterization of a new Catechol 2,3-dioxygenase isolated from a Gram-positive bacterium able to utilize phenol as the sole carbon and energy source. Planococcus sp. strain S5 grown on 1 or 2 mM phenol showed activity of both a Catechol 1,2- and Catechol 2,3-dioxygenase while at a higher concentrations of phenol only Catechol 2,3-dioxygenase activity was observed. The enzyme was optimally active at 60°C and pH 8.0. Kinetic studies showed that the K(m) and V(max) of the enzyme were 42.70 µM and 329.96 mU, respectively. The Catechol 2,3-dioxygenase showed the following relative meta-cleavage activities for various Catechols tested: Catechol (100%), 3-methylCatechol (13.67%), 4-methylCatechol (106.33%) and 4-chloroCatechol (203.80%). The high reactivity of this enzyme towards 4-chloroCatechol is different from that observed for other Catechol 2,3-dioxygenases. Nucleotide sequencing and homology search revealed that the gene encoding the S5 Catechol 2,3-dioxygenase shared the greatest homology with the known genes encoding isoenzymes from Gram-negative Pseudomonas strains.
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properties of Catechol 2 3 dioxygenase from crude extract of stenotrophomonas maltophilia strain kb2 immobilized in calcium alginate hydrogels
Biochemical Engineering Journal, 2012Co-Authors: Danuta Wojcieszynska, Katarzyna Hupertkocurek, Anna Jankowska, Urszula GuzikAbstract:Abstract In this paper we report the immobilization of Catechol 2,3-dioxygenase from Stenotrophomonas maltophilia KB2 in alginate hydrogel with the aim of improving its functional stability by increasing structural rigidity of the enzyme. Immobilization yield and expressed activity were 49.4% and 49.4%, respectively. The storage stability of entrapped Catechol 2,3-dioxygenase at 4 °C was found up to 35 days (266.3 mU/mg protein), while at 4 °C the free enzyme lost its activity within 24 h. Immobilization of dioxygenase increased the optimum temperature for activity by 10 °C, while both soluble and immobilized enzyme showed maximum activity at the same pH. The Km, Vmax, and Hill constant values for immobilized enzyme were 0.2 μM, 604.6 mU/mg protein, and 1.00, respectively, whereas those for the free enzyme were 46.3 μM, 1602.0 mU/mg protein, and 4.1, respectively.The immobilized Catechol 2,3-dioxygenase from KB2 strain showed relatively higher activity against 3-methylCatechol, 4-methylCatechol, 4,5-dichloroCatechol, 3,5-dichloroCatechol, hydroquinone and tetrachlorohydroquinone than soluble enzyme. Immobilization of Catechol 2,3-dioxygenase from KB2 strain protected the enzyme from the inhibition and enhanced its resistance to inactivation during catalysis. That makes the enzyme suitable for the bioremediation and detoxification of xenobiotic-contaminated environments.
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high activity Catechol 2 3 dioxygenase from the cresols degrading stenotrophomonas maltophilia strain kb2
International Biodeterioration & Biodegradation, 2011Co-Authors: Danuta Wojcieszynska, Katarzyna Hupertkocurek, Izabela Gren, Urszula GuzikAbstract:Abstract This study aimed at characterization of Catechol 2,3-dioxygenase from Stenotrophomonas maltophilia KB2, being able to utilize a wide spectrum of aromatic substrates as a sole carbon and energy source. 2-methylphenol, 3-methylphenol, and 4-methylphenol was completely degraded during 24 h in concentration 6 mM, 7 mM, and 5 mM, respectively. When cells of strain KB2 were growing on methylphenols, Catechol 2,3-dioxygenase was induced. Biochemical analysis revealed that the examined enzyme was similar to another Catechol 2,3-dioxygenases, but showed extremely high activity. The enzyme was optimally active at 30 °C and pH 7.6. Kinetic studies showed that the value of K m , V max and Hill constant was 85.11 μM, 3.08 μM min −1 and 4.09 respectively. Comparative structural and phylogenetic analysis of Catechol 2,3-dioxygenase from S. maltophilia KB2 had placed the protein with the single-ring substrate subfamily of the extradiol dioxygenase. We observed the presence of externally located α-helices and internally located β-sheets. We also suggest that the Fe 2+ ion binding is facilitated via four ligands: two histidine residues, one glutamate residue and one molecule of water.
Danuta Wojcieszynska - One of the best experts on this subject based on the ideXlab platform.
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characterization of Catechol 2 3 dioxygenase from planococcus sp strain s5 induced by high phenol concentration
Acta Biochimica Polonica, 2012Co-Authors: Katarzyna Hupertkocurek, Urszula Guzik, Danuta WojcieszynskaAbstract:This study aimed at characterization of a new Catechol 2,3-dioxygenase isolated from a Gram-positive bacterium able to utilize phenol as the sole carbon and energy source. Planococcus sp. strain S5 grown on 1 or 2 mM phenol showed activity of both a Catechol 1,2- and Catechol 2,3-dioxygenase while at a higher concentrations of phenol only Catechol 2,3-dioxygenase activity was observed. The enzyme was optimally active at 60°C and pH 8.0. Kinetic studies showed that the K(m) and V(max) of the enzyme were 42.70 µM and 329.96 mU, respectively. The Catechol 2,3-dioxygenase showed the following relative meta-cleavage activities for various Catechols tested: Catechol (100%), 3-methylCatechol (13.67%), 4-methylCatechol (106.33%) and 4-chloroCatechol (203.80%). The high reactivity of this enzyme towards 4-chloroCatechol is different from that observed for other Catechol 2,3-dioxygenases. Nucleotide sequencing and homology search revealed that the gene encoding the S5 Catechol 2,3-dioxygenase shared the greatest homology with the known genes encoding isoenzymes from Gram-negative Pseudomonas strains.
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properties of Catechol 2 3 dioxygenase from crude extract of stenotrophomonas maltophilia strain kb2 immobilized in calcium alginate hydrogels
Biochemical Engineering Journal, 2012Co-Authors: Danuta Wojcieszynska, Katarzyna Hupertkocurek, Anna Jankowska, Urszula GuzikAbstract:Abstract In this paper we report the immobilization of Catechol 2,3-dioxygenase from Stenotrophomonas maltophilia KB2 in alginate hydrogel with the aim of improving its functional stability by increasing structural rigidity of the enzyme. Immobilization yield and expressed activity were 49.4% and 49.4%, respectively. The storage stability of entrapped Catechol 2,3-dioxygenase at 4 °C was found up to 35 days (266.3 mU/mg protein), while at 4 °C the free enzyme lost its activity within 24 h. Immobilization of dioxygenase increased the optimum temperature for activity by 10 °C, while both soluble and immobilized enzyme showed maximum activity at the same pH. The Km, Vmax, and Hill constant values for immobilized enzyme were 0.2 μM, 604.6 mU/mg protein, and 1.00, respectively, whereas those for the free enzyme were 46.3 μM, 1602.0 mU/mg protein, and 4.1, respectively.The immobilized Catechol 2,3-dioxygenase from KB2 strain showed relatively higher activity against 3-methylCatechol, 4-methylCatechol, 4,5-dichloroCatechol, 3,5-dichloroCatechol, hydroquinone and tetrachlorohydroquinone than soluble enzyme. Immobilization of Catechol 2,3-dioxygenase from KB2 strain protected the enzyme from the inhibition and enhanced its resistance to inactivation during catalysis. That makes the enzyme suitable for the bioremediation and detoxification of xenobiotic-contaminated environments.
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high activity Catechol 2 3 dioxygenase from the cresols degrading stenotrophomonas maltophilia strain kb2
International Biodeterioration & Biodegradation, 2011Co-Authors: Danuta Wojcieszynska, Katarzyna Hupertkocurek, Izabela Gren, Urszula GuzikAbstract:Abstract This study aimed at characterization of Catechol 2,3-dioxygenase from Stenotrophomonas maltophilia KB2, being able to utilize a wide spectrum of aromatic substrates as a sole carbon and energy source. 2-methylphenol, 3-methylphenol, and 4-methylphenol was completely degraded during 24 h in concentration 6 mM, 7 mM, and 5 mM, respectively. When cells of strain KB2 were growing on methylphenols, Catechol 2,3-dioxygenase was induced. Biochemical analysis revealed that the examined enzyme was similar to another Catechol 2,3-dioxygenases, but showed extremely high activity. The enzyme was optimally active at 30 °C and pH 7.6. Kinetic studies showed that the value of K m , V max and Hill constant was 85.11 μM, 3.08 μM min −1 and 4.09 respectively. Comparative structural and phylogenetic analysis of Catechol 2,3-dioxygenase from S. maltophilia KB2 had placed the protein with the single-ring substrate subfamily of the extradiol dioxygenase. We observed the presence of externally located α-helices and internally located β-sheets. We also suggest that the Fe 2+ ion binding is facilitated via four ligands: two histidine residues, one glutamate residue and one molecule of water.
Hang Min - One of the best experts on this subject based on the ideXlab platform.
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Isolation and characterization of phenanthrene-degrading Sphingomonas paucimobilis strain ZX4.
Biodegradation, 2005Co-Authors: Ying Xia, Hang Min, Gang Rao, Ji Liu, Xue-jun DuanAbstract:Phenanthrene-degrading bacterium strain ZX4 was isolated from an oil-contaminated soil, and identified as Sphingomonaspaucimobilis based on 16S rDNA sequence, cellular fatty acid composition, mol% G + C and Biolog-GN tests. Besides phenanthrene, strain ZX4 could also utilize naphthalene, fluorene and other aromatic compounds. The growth on salicylic acid and Catechol showed that the strain degraded phenanthrene via salicylate pathway, while the assay of Catechol 2, 3-dioxygenase revealed Catechol could be metabolized through meta-cleavage pathway. Three genes, including two of meta-cleavage operon genes and one of GST encoding gene were obtained. The order of genes arrangement was similar to S-type meta-pathway operons. The phylogenetic trees based on 16S rDNA sequence and meta-pathway gene both revealed that strain ZX4 is clustered with strains from genus Sphingomonas.
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Characterization and phylogenetic analysis of a phenanthrene-degrading strain isolated from oil-contaminated soil.
Journal of environmental sciences (China), 2004Co-Authors: Ying Xia, Hang MinAbstract:Bacterium strain EVA17 was isolated from an oil-contaminated soil, and identified as Sphingomonas sp. based on analysis of 16S rDNA sequence, cellular fatty acid composition and physiological-chemical tests. The salicylate hydroxylase and Catechol 2, 3-dioxygenase (C230) were detected in cell-free lysates, suggesting a pathway for phenanthrene catabolism via salicylate and Catechol. Alignment showed that both of the C230 and GST genes of the strain EVA17 had high similarity with homologues of strains from genus Sphingomonas. The phylogenetic analysis based on 16S rDNA and C230 gene sequence indicated that EVA17 should be classified into genus Sphingomonas, although the two phylogenetic trees were slightly different from each other. The results of coamplification and sequence determination indicated that GST gene should be located upstream of the C230 gene.
Katarzyna Hupertkocurek - One of the best experts on this subject based on the ideXlab platform.
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characterization of Catechol 2 3 dioxygenase from planococcus sp strain s5 induced by high phenol concentration
Acta Biochimica Polonica, 2012Co-Authors: Katarzyna Hupertkocurek, Urszula Guzik, Danuta WojcieszynskaAbstract:This study aimed at characterization of a new Catechol 2,3-dioxygenase isolated from a Gram-positive bacterium able to utilize phenol as the sole carbon and energy source. Planococcus sp. strain S5 grown on 1 or 2 mM phenol showed activity of both a Catechol 1,2- and Catechol 2,3-dioxygenase while at a higher concentrations of phenol only Catechol 2,3-dioxygenase activity was observed. The enzyme was optimally active at 60°C and pH 8.0. Kinetic studies showed that the K(m) and V(max) of the enzyme were 42.70 µM and 329.96 mU, respectively. The Catechol 2,3-dioxygenase showed the following relative meta-cleavage activities for various Catechols tested: Catechol (100%), 3-methylCatechol (13.67%), 4-methylCatechol (106.33%) and 4-chloroCatechol (203.80%). The high reactivity of this enzyme towards 4-chloroCatechol is different from that observed for other Catechol 2,3-dioxygenases. Nucleotide sequencing and homology search revealed that the gene encoding the S5 Catechol 2,3-dioxygenase shared the greatest homology with the known genes encoding isoenzymes from Gram-negative Pseudomonas strains.
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properties of Catechol 2 3 dioxygenase from crude extract of stenotrophomonas maltophilia strain kb2 immobilized in calcium alginate hydrogels
Biochemical Engineering Journal, 2012Co-Authors: Danuta Wojcieszynska, Katarzyna Hupertkocurek, Anna Jankowska, Urszula GuzikAbstract:Abstract In this paper we report the immobilization of Catechol 2,3-dioxygenase from Stenotrophomonas maltophilia KB2 in alginate hydrogel with the aim of improving its functional stability by increasing structural rigidity of the enzyme. Immobilization yield and expressed activity were 49.4% and 49.4%, respectively. The storage stability of entrapped Catechol 2,3-dioxygenase at 4 °C was found up to 35 days (266.3 mU/mg protein), while at 4 °C the free enzyme lost its activity within 24 h. Immobilization of dioxygenase increased the optimum temperature for activity by 10 °C, while both soluble and immobilized enzyme showed maximum activity at the same pH. The Km, Vmax, and Hill constant values for immobilized enzyme were 0.2 μM, 604.6 mU/mg protein, and 1.00, respectively, whereas those for the free enzyme were 46.3 μM, 1602.0 mU/mg protein, and 4.1, respectively.The immobilized Catechol 2,3-dioxygenase from KB2 strain showed relatively higher activity against 3-methylCatechol, 4-methylCatechol, 4,5-dichloroCatechol, 3,5-dichloroCatechol, hydroquinone and tetrachlorohydroquinone than soluble enzyme. Immobilization of Catechol 2,3-dioxygenase from KB2 strain protected the enzyme from the inhibition and enhanced its resistance to inactivation during catalysis. That makes the enzyme suitable for the bioremediation and detoxification of xenobiotic-contaminated environments.
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high activity Catechol 2 3 dioxygenase from the cresols degrading stenotrophomonas maltophilia strain kb2
International Biodeterioration & Biodegradation, 2011Co-Authors: Danuta Wojcieszynska, Katarzyna Hupertkocurek, Izabela Gren, Urszula GuzikAbstract:Abstract This study aimed at characterization of Catechol 2,3-dioxygenase from Stenotrophomonas maltophilia KB2, being able to utilize a wide spectrum of aromatic substrates as a sole carbon and energy source. 2-methylphenol, 3-methylphenol, and 4-methylphenol was completely degraded during 24 h in concentration 6 mM, 7 mM, and 5 mM, respectively. When cells of strain KB2 were growing on methylphenols, Catechol 2,3-dioxygenase was induced. Biochemical analysis revealed that the examined enzyme was similar to another Catechol 2,3-dioxygenases, but showed extremely high activity. The enzyme was optimally active at 30 °C and pH 7.6. Kinetic studies showed that the value of K m , V max and Hill constant was 85.11 μM, 3.08 μM min −1 and 4.09 respectively. Comparative structural and phylogenetic analysis of Catechol 2,3-dioxygenase from S. maltophilia KB2 had placed the protein with the single-ring substrate subfamily of the extradiol dioxygenase. We observed the presence of externally located α-helices and internally located β-sheets. We also suggest that the Fe 2+ ion binding is facilitated via four ligands: two histidine residues, one glutamate residue and one molecule of water.
Yves Jouanneau - One of the best experts on this subject based on the ideXlab platform.
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a novel 2fe 2s ferredoxin from pseudomonas putidamt2 promotes the reductive reactivation of Catechol 2 3 dioxygenase
Journal of Biological Chemistry, 1998Co-Authors: N Hugo, Jean Armengaud, Jacques Gaillard, Kenneth N Timmis, Yves JouanneauAbstract:Catechol 2,3-dioxygenase (XylE) is a component of the TOL plasmid-encoded pathway for the degradation of toluene and xylenes and catalyzes the dioxygenolytic cleavage of the aromatic ring. Purified XylE is oxygen-sensitive and unstable in vitro, particularly in the presence of substituted Catechol substrates, but it is stabilized in vivo by another protein, XylT, encoded by the xylT gene located just upstream of xylE. In this study, we have purified to homogeneity the XylT product from a recombinant Escherichia coli strain containing a hyperexpressible xylT gene and characterized it as a novel [2Fe-2S] ferredoxin. It is the first example of a soluble ferredoxin with a net positive charge at neutral pH. The EPR signal of the iron sulfur cluster has rhombic symmetry as is the case for plant-type ferredoxins, but the XylT absorbance spectrum resembles more closely that of adrenodoxin. The midpoint redox potential was determined to be −373 ± 6 mV, at pH 8.5. XylT was unusually unstable for a [2Fe-2S] ferredoxin, with half-lives of 69 min at 25 °C in air and 70 min at 37 °C in argon. With photochemically reduced 5-deazaflavin for the controlled generation of reductant, it was demonstrated that XylT mediates the rapid reactivation of purified inactive Catechol 2,3-dioxygenase in vitro. Inactivation of XylE by 4-methylCatechol resulted in oxidation of the active site iron to a high spin ferric state that was detectable by EPR. Spectroscopic evidence presented here demonstrates that XylT reactivates XylE through reduction of the iron atom in the active site of the enzyme. It is the first instance of a ferredoxin-mediated reactivation of an enzyme. The level of expression of XylT in Pseudomonas putida mt2 cells is low and the calculated XylT/XylE molar ratio is consistent with the proposal that XylE reactivation involves catalytic nonstoichiometric amounts of XylT.