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Lawrence P Wackett - One of the best experts on this subject based on the ideXlab platform.
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Toluene 2-Monooxygenase-Dependent Growth of Burkholderia cepacia G4/PR1 on Diethyl Ether.
Applied and Environmental Microbiology, 1997Co-Authors: Lisa M. Newman, Lawrence P Wackett, Michael J. SadowskyAbstract:Aerobic bacterial growth on aromatic hydrocarbons typically requires oxygenase enzymes, which are known to fortuitously oxidize nongrowth substrates. In this study, we found that oxidation of diethyl ether by Toluene 2-monooxygenase supported more rapid growth of Burkholderia cepacia G4/PR1 than did the aromatic substrates n-propylbenzene and o-xylene. The wild-type Burkholderia cepacia G4 failed to grow on diethyl ether. Purified Toluene 2-monooxygenase protein components oxidized diethyl ether stoichiometrically to ethanol and acetaldehyde. Butyl methyl ether, diethyl sulfide, and 2-chloroethyl ethyl ether were oxidized by B. cepacia G4/PR1.
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Trichloroethylene oxidation by purified Toluene 2-monooxygenase: products, kinetics, and turnover-dependent inactivation.
Journal of bacteriology, 1997Co-Authors: Lisa M. Newman, Lawrence P WackettAbstract:Trichloroethylene is oxidized by several types of nonspecific bacterial oxygenases. Toluene 2-monooxygenase from Burkholderia cepacia G4 is implicated in trichloroethylene oxidation and is uniquely suggested to be resistant to turnover-dependent inactivation in vivo. In this work, the oxidation of trichloroethylene was studied with purified Toluene 2-monooxygenase. All three purified Toluene 2-monooxygenase protein components and NADH were required to reconstitute full trichloroethylene oxidation activity in vitro. The apparent Km and Vmax were 12 microM and 37 nmol per min per mg of hydroxylase component, respectively. Ten percent of the full activity was obtained when the small-molecular-weight enzyme component was omitted. The stable oxidation products, accounting for 84% of the trichloroethylene oxidized, were carbon monoxide, formic acid, glyoxylic acid, and covalently modified oxygenase proteins that constituted 12% of the reacted [14C]trichloroethylene. The stable oxidation products may all derive from the unstable intermediate trichloroethylene epoxide that was trapped by reaction with 4-(p-nitrobenzyl)pyridine. Chloral hydrate and dichloroacetic acid were not detected. This finding differs from that with soluble methane monooxygenase and cytochrome P-450 monooxygenase, which produce chloral hydrate. Trichloroethylene-dependent inactivation of Toluene 2-monooxygenase activity was observed. All of the protein components were covalently modified during the oxidation of trichloroethylene. The addition of cysteine to reaction mixtures partially protected the enzyme system against inactivation, most notably protecting the NADH-oxidoreductase component. This suggested the participation of diffusible intermediates in the inactivation of the oxidoreductase.
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PURIFICATION AND CHARACTERIZATION OF Toluene 2-MONOOXYGENASE FROM BURKHOLDERIA CEPACIA G4
Biochemistry, 1995Co-Authors: Lisa M. Newman, Lawrence P WackettAbstract:Recent in vivo studies indicate that ring monooxygenation is a widespread mechanism by which bacteria metabolize aromatic hydrocarbons and obtain carbon and energy. In this study, Toluene 2-monooxygenase from Burkholderia (formerly Pseudomonas) cepacia G4 was purified to homogeneity and found to be a three-component enzyme system. The reconstituted enzyme system oxidized Toluene to o-cresol and o-cresol to 3-methylcatechol, an important intermediate for growth of the bacterium on Toluene. Steady-state kinetic parameters measured for the water-soluble substrate o-cresol were a Km of 0.8 microM and a Vmax of 131 nmol min-1 (mg of hydroxylase protein)-1. The three protein components were (1) a 40 kDa polypeptide containing one FAD and a [2Fe2S] cluster, (2) a 10.4 kDa polypeptide that contained no identifiable metals or organic cofactors, and (3) a 211 kDa alpha 2 beta 2 gamma 2 component containing five to six iron atoms. The 40 kDa flavo-iron-sulfur protein oxidized NADH and transferred electrons to cytochrome c, dyes, and the alpha 2 beta 2 gamma 2 component. It is analogous to other NADH oxidoreductase components found in a wide range of bacterial mono- and dioxygenases. The 10.4 kDa component, added to the other two components and NADH, increased Toluene oxidation rates 10-fold. The alpha 2 beta 2 gamma 2 component was indicated to contain the site for Toluene binding and hydroxylation by the following observations: (1) tight binding to a Toluene affinity column; (2) oxidation of Toluene after reduction of the protein with dithionite and adding O2; (3) H2O2-dependent Toluene oxidation and catalase activity; and (4) spectroscopic studies of the iron atoms in the component. The alpha 2 beta 2 gamma 2 component had no significant absorbance in the visible region. EPR spectroscopy yielded a signal at g = 16 upon addition of > 2 equiv of electrons per 2 Fe atoms. Taken with the quantitation of five to six iron atoms, the data suggest that the alpha 2 beta 2 gamma 2 component contains two binuclear iron centers. In total, the structural, spectroscopic, and catalytic features of Toluene 2-monooxygenase are reminiscent of soluble methane monooxygenase obtained from methanotrophic bacteria. The two enzyme systems also differ in many subtle ways; for example, they oxidize Toluene with completely different regiospecificity.
Lisa M. Newman - One of the best experts on this subject based on the ideXlab platform.
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Toluene 2-Monooxygenase-Dependent Growth of Burkholderia cepacia G4/PR1 on Diethyl Ether.
Applied and Environmental Microbiology, 1997Co-Authors: Lisa M. Newman, Lawrence P Wackett, Michael J. SadowskyAbstract:Aerobic bacterial growth on aromatic hydrocarbons typically requires oxygenase enzymes, which are known to fortuitously oxidize nongrowth substrates. In this study, we found that oxidation of diethyl ether by Toluene 2-monooxygenase supported more rapid growth of Burkholderia cepacia G4/PR1 than did the aromatic substrates n-propylbenzene and o-xylene. The wild-type Burkholderia cepacia G4 failed to grow on diethyl ether. Purified Toluene 2-monooxygenase protein components oxidized diethyl ether stoichiometrically to ethanol and acetaldehyde. Butyl methyl ether, diethyl sulfide, and 2-chloroethyl ethyl ether were oxidized by B. cepacia G4/PR1.
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Trichloroethylene oxidation by purified Toluene 2-monooxygenase: products, kinetics, and turnover-dependent inactivation.
Journal of bacteriology, 1997Co-Authors: Lisa M. Newman, Lawrence P WackettAbstract:Trichloroethylene is oxidized by several types of nonspecific bacterial oxygenases. Toluene 2-monooxygenase from Burkholderia cepacia G4 is implicated in trichloroethylene oxidation and is uniquely suggested to be resistant to turnover-dependent inactivation in vivo. In this work, the oxidation of trichloroethylene was studied with purified Toluene 2-monooxygenase. All three purified Toluene 2-monooxygenase protein components and NADH were required to reconstitute full trichloroethylene oxidation activity in vitro. The apparent Km and Vmax were 12 microM and 37 nmol per min per mg of hydroxylase component, respectively. Ten percent of the full activity was obtained when the small-molecular-weight enzyme component was omitted. The stable oxidation products, accounting for 84% of the trichloroethylene oxidized, were carbon monoxide, formic acid, glyoxylic acid, and covalently modified oxygenase proteins that constituted 12% of the reacted [14C]trichloroethylene. The stable oxidation products may all derive from the unstable intermediate trichloroethylene epoxide that was trapped by reaction with 4-(p-nitrobenzyl)pyridine. Chloral hydrate and dichloroacetic acid were not detected. This finding differs from that with soluble methane monooxygenase and cytochrome P-450 monooxygenase, which produce chloral hydrate. Trichloroethylene-dependent inactivation of Toluene 2-monooxygenase activity was observed. All of the protein components were covalently modified during the oxidation of trichloroethylene. The addition of cysteine to reaction mixtures partially protected the enzyme system against inactivation, most notably protecting the NADH-oxidoreductase component. This suggested the participation of diffusible intermediates in the inactivation of the oxidoreductase.
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PURIFICATION AND CHARACTERIZATION OF Toluene 2-MONOOXYGENASE FROM BURKHOLDERIA CEPACIA G4
Biochemistry, 1995Co-Authors: Lisa M. Newman, Lawrence P WackettAbstract:Recent in vivo studies indicate that ring monooxygenation is a widespread mechanism by which bacteria metabolize aromatic hydrocarbons and obtain carbon and energy. In this study, Toluene 2-monooxygenase from Burkholderia (formerly Pseudomonas) cepacia G4 was purified to homogeneity and found to be a three-component enzyme system. The reconstituted enzyme system oxidized Toluene to o-cresol and o-cresol to 3-methylcatechol, an important intermediate for growth of the bacterium on Toluene. Steady-state kinetic parameters measured for the water-soluble substrate o-cresol were a Km of 0.8 microM and a Vmax of 131 nmol min-1 (mg of hydroxylase protein)-1. The three protein components were (1) a 40 kDa polypeptide containing one FAD and a [2Fe2S] cluster, (2) a 10.4 kDa polypeptide that contained no identifiable metals or organic cofactors, and (3) a 211 kDa alpha 2 beta 2 gamma 2 component containing five to six iron atoms. The 40 kDa flavo-iron-sulfur protein oxidized NADH and transferred electrons to cytochrome c, dyes, and the alpha 2 beta 2 gamma 2 component. It is analogous to other NADH oxidoreductase components found in a wide range of bacterial mono- and dioxygenases. The 10.4 kDa component, added to the other two components and NADH, increased Toluene oxidation rates 10-fold. The alpha 2 beta 2 gamma 2 component was indicated to contain the site for Toluene binding and hydroxylation by the following observations: (1) tight binding to a Toluene affinity column; (2) oxidation of Toluene after reduction of the protein with dithionite and adding O2; (3) H2O2-dependent Toluene oxidation and catalase activity; and (4) spectroscopic studies of the iron atoms in the component. The alpha 2 beta 2 gamma 2 component had no significant absorbance in the visible region. EPR spectroscopy yielded a signal at g = 16 upon addition of > 2 equiv of electrons per 2 Fe atoms. Taken with the quantitation of five to six iron atoms, the data suggest that the alpha 2 beta 2 gamma 2 component contains two binuclear iron centers. In total, the structural, spectroscopic, and catalytic features of Toluene 2-monooxygenase are reminiscent of soluble methane monooxygenase obtained from methanotrophic bacteria. The two enzyme systems also differ in many subtle ways; for example, they oxidize Toluene with completely different regiospecificity.
Yanji Wang - One of the best experts on this subject based on the ideXlab platform.
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direct synthesis of dimethyl Toluene 2 4 dicarbamate from 2 4 Toluene diamine urea and methanol
Industrial & Engineering Chemistry Research, 2011Co-Authors: Xinqiang Zhao, Na Wang, Yanlou Geng, Yanji WangAbstract:Dimethyl Toluene-2,4-dicarbamate (TDC) was directly synthesized from 2,4-Toluene diamine (TDA), urea, and methanol in order to overcome the drawbacks of other technological routes to TDC. First the thermodynamic analysis for this reaction was made and the results show that the reaction is endothermic and can occur spontaneously beyond 413.8 K. Then the effects of catalyst and reaction conditions were studied. TDA conversion of 98.8% and TDC selectivity of 41.6% were attained in the presence of zinc chloride catalyst and under the suitable conditions of molar ratio of TDA/zinc chloride/urea/methanol = 1/0.07/5/80, reaction temperature of 190 °C, reaction pressure of 3.0 MPa, and reaction time of 9 h. Low TDC selectivity is attributed to the difficulty in the conversion of the intermediates, methyl 2-methyl-5-amino N-phenylcarbamate (TMC1) and methyl 3-amino-4-methyl-N-phenylcarbamate (TMC2), to TDC. Finally on the basis of analyses of HPLC–MS, HPLC, and GC, three possible reaction paths were proposed. One ...
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catalytic synthesis of Toluene 2 4 diisocyanate from dimethyl carbonate
Journal of Chemical Technology & Biotechnology, 2001Co-Authors: Yanji Wang, Xinqiang Zhao, Shufang Wang, Jiyan ZhangAbstract:The process for catalytic synthesis of Toluene-2,4-diisocyanate (TDI) from dimethyl carbonate (DMC) consists of two steps. Starting from the catalytic reaction between Toluene-2,4-diamine (TDA) and DMC, dimethyl Toluene-2,4-dicarbamate (TDC) is formed, and then decomposed to TDI. For the first step, the yield of TDC is 53.5% at a temperature of 250 °C, over Zn(OAc)2/α–Al2O3 catalyst. For the second step, the yield of TDI is 92.6% at temperatures of 250–270 °C and under pressure of 2.7 kPa, over uranyl zinc acetate catalyst, when di-n-octyl sebacate(DOS) is used as heat-carrier, and a mixture of tetrahydrofuran (THF) and nitrobenzene is used as solvent. © 2001 Society of Chemical Industry
Rui Fausto - One of the best experts on this subject based on the ideXlab platform.
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Synthesis of Thieno[2,3-d]pyrimidines via Microwave-Assisted Thermolysis of 1-(Thiophene-2-yl)-1H-tetrazoles
ChemistrySelect, 2016Co-Authors: Maria I. L. Soares, Teresa M. V. D. Pinho E Melo, Rui FaustoAbstract:A selective and efficient method for the synthesis of unusual thieno[2,3-d]pyrimidines incorporating two thiophene rings via thermal reactions of 1-(thiophene-2-yl)-1H-tetrazoles is reported. Microwave-assisted thermolysis of 1-(thiophene-2-yl)-1H-tetrazoles at 160 °C in a water/Toluene (2:1) solvent system afforded thieno[2,3-d]pyrimidines in moderate to good yield (up to 66 %) as single products.
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Synthesis of Thieno[2,3‐d]pyrimidines via Microwave‐Assisted Thermolysis of 1‐(Thiophene‐2‐yl)‐1H‐tetrazoles
ChemistrySelect, 2016Co-Authors: Maria I. L. Soares, Teresa M. V. D. Pinho E Melo, Rui FaustoAbstract:A selective and efficient method for the synthesis of unusual thieno[2,3-d]pyrimidines incorporating two thiophene rings via thermal reactions of 1-(thiophene-2-yl)-1H-tetrazoles is reported. Microwave-assisted thermolysis of 1-(thiophene-2-yl)-1H-tetrazoles at 160 °C in a water/Toluene (2:1) solvent system afforded thieno[2,3-d]pyrimidines in moderate to good yield (up to 66 %) as single products.
Yan-ping Chen - One of the best experts on this subject based on the ideXlab platform.
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Solid−Liquid Equilibria for 3-Aminophenol + 2-Aminophenol, 4-(Acetylamino)Toluene + 2-(Acetylamino)Toluene, and 3,4,5-Trimethoxybenzoic Acid + 2,4,5-Trimethoxybenzoic Acid
Journal of Chemical & Engineering Data, 2005Co-Authors: Chih-yung Huang, Muoi Tang, Yan-ping ChenAbstract:Solid−liquid equilibria for three isomeric binary mixtures of 3-aminophenol (1) + 2-aminophenol (2) (eutectic temperature TE = 376.5 K, eutectic composition x1E = 0.86), 4-(acetylamino)Toluene (1) + 2-(acetylamino)Toluene (2) (TE = 357.2 K, x1E = 0.30), and 3,4,5-trimethoxybenzoic acid (1) + 2,4,5-trimethoxybenzoic acid (2) (TE = 395.8 K, x1E = 0.33) were measured using differential scanning calorimetry (DSC). Simple eutectic behaviors for these systems were observed. The experimental results were correlated using the Wilson activity coefficient model. The eutectic compositions were also determined using Gray's model and the fractional transformation method.