Biodegradation of cyclic amines by aPseudomonasstrain involves an amine mono-oxygenase

2003 ◽  
Vol 49 (3) ◽  
pp. 181-188 ◽  
Author(s):  
M Trigui ◽  
S Pulvin ◽  
P Poupin ◽  
D Thomas
Keyword(s):  
Pseudomonas Putida ◽  
Specific Inhibitor ◽  
Sole Source ◽  
Spectrophotometric Analysis ◽  
Piperidine Ring ◽  
Ring Cleavage ◽  
Cyclic Amines ◽  
Growth Substrates

Pseudomonas putida O1G3 catalyzes the degradation of pyrrolidine and piperidine. This strain can use these compounds as the sole source of carbon, nitrogen, and energy. When the cyclic amines were used as the growth substrates, the synthesis of a soluble heme amine mono-oxygenase was induced in this bacteria. This observation was confirmed by spectrophotometric analysis and specific inhibitor. This mono-oxygenase is a NADH-dependent enzyme and catalyzes the cleavage of the C—N bond of the pyrrolidine and piperidine ring by a mechanism similar to a N dealkylation. This reaction could be followed by ring cleavage to form γ-aminobutyraldehyde oxidized to γ-aminobutyrate. Further investigations to purify the heme-containing mono-oxygenase are in progress.Key words: biodegradation, Pseudomonas, pyrrolidine, piperidine, amine mono-oxygenase.

scholarly journals Degradation of Morpholine by an EnvironmentalMycobacterium Strain Involves a Cytochrome P-450

1998 ◽  
Vol 64 (1) ◽  
pp. 159-165 ◽  
Author(s):  
P. Poupin ◽  
N. Truffaut ◽  
B. Combourieu ◽  
P. Besse ◽  
M. Sancelme ◽  
...  
Keyword(s):  
Specific Inhibitor ◽  
Direct Analysis ◽  
Difference Spectrum ◽  
Sole Source ◽  
Spectrophotometric Analysis ◽  
Treatment Unit ◽  
H Nmr ◽  
Cell Extracts ◽  
Cytochrome P 450

A Mycobacterium strain (RP1) was isolated from a contaminated activated sludge collected in a wastewater treatment unit of a chemical plant. It was capable of utilizing morpholine and other heterocyclic compounds, such as pyrrolidine and piperidine, as the sole source of carbon, nitrogen, and energy. The use of in situ1H nuclear magnetic resonance (1H NMR) spectroscopy allowed the determination of two intermediates in the biodegradative pathway, 2-(2-aminoethoxy)acetate and glycolate. The inhibitory effects of metyrapone on the degradative abilities of strain RP1 indicated the involvement of a cytochrome P-450 in the biodegradation of morpholine. This observation was confirmed by spectrophotometric analysis and1H NMR. Reduced cell extracts from morpholine-grown cultures, but not succinate-grown cultures, gave rise to a carbon monoxide difference spectrum with a peak near 450 nm, which indicated the presence of a soluble cytochrome P-450. 1H NMR allowed the direct analysis of the incubation medium containing metyrapone, a specific inhibitor of cytochrome P-450. The inhibition of morpholine degradation was dependent on the morpholine/metyrapone ratio. The heme-containing monooxygenase was also detected in pyrrolidine- and piperidine-grown cultures. The abilities of different compounds to support strain growth or the induction of a soluble cytochrome P-450 were assayed. The results suggest that this enzyme catalyzes the cleavage of the C—N bond of the morpholine ring.

Synthesis of Medium-Sized Cyclic Amines by Selective Ring Cleavage of Sulfonylated Bicyclic Amines

2001 ◽  
Vol 3 (19) ◽  
pp. 2957-2960 ◽  
Author(s):  
Fátima Iradier ◽  
Ramón Gómez Arrayás ◽  
Juan Carlos Carretero
Keyword(s):  
Ring Cleavage ◽  
Cyclic Amines

Differential Roles of Three Different Upper Pathway Meta Ring Cleavage Product Hydrolases in the Degradation of Dibenzo- p -dioxin and Dibenzofuran by Sphingomonas wittichii RW1

2021 ◽  
Author(s):  
Thamer Y. Mutter ◽  
Gerben J. Zylstra
Keyword(s):  
Gene Knockout ◽  
Sole Source ◽  
Cleavage Product ◽  
Ring Cleavage ◽  
Catabolic Pathway ◽  
Double Knockout ◽  
Cleavage Enzyme ◽  
Sphingomonas Wittichii ◽  
Physiological Approach

Sphingomonas wittichii RW1 grows on the two related compounds dibenzofuran (DBF) and dibenzo- p -dioxin (DXN) as the sole source of carbon. Previous work by others (P.V. Bunz, R. Falchetto, and A.M. Cook. Biodegradation 4:171-8, 1993, doi: 10.1007/BF00695119) identified two upper pathway meta cleavage product hydrolases (DxnB1 and DxnB2) active on the DBF upper pathway metabolite 2-hydroxy-6-oxo-6-(2-hydroxyphenyl)-hexa-2,4-dienoate. We took a physiological approach to determine the role of these two enzymes in the degradation of DBF and DXN by RW1. Single knockouts of either plasmid located dbfB1 or chromosome located dbfB2 had no effect on RW1 growth on either DBF or DXN. However, a double knockout lost the ability to grow on DBF but still grew normally on DXN demonstrating that DbfB1 and DbfB2 are the only hydrolases involved in the DBF upper pathway. Using a transcriptomic-guided approach we identified a constitutively expressed third hydrolase encoded by the chromosomally located SWIT0910 gene. Knockout of SWIT0910 resulted in a strain that no longer grows on DXN but still grows normally on DBF. Thus the DbfB1 and DbfB2 hydrolases function in the DBF but not the DXN catabolic pathway and the SWIT0190 hydrolase functions in the DXN but not the DBF catabolic pathway. Importance S. wittichii RW1 is one of only a few strains known to grow on DXN as the sole course of carbon. Much of the work deciphering the related RW1 DXN and DBF catabolic pathways has involved genome gazing, transcriptomics, proteomics, heterologous expression, and enzyme purification and characterization. Very little research has utilized physiological techniques to precisely dissect the genes and enzymes involved in DBF and DXN degradation. Previous work by others identified and extensively characterized two RW1 upper pathway hydrolases. Our present work demonstrates that these two enzymes are involved in DBF but not DXN degradation. In addition, our work identified a third constitutively expressed hydrolase that is involved in DXN but not DBF degradation. Combined with our previous work, this means that the RW1 DXN upper pathway involves genes from three very different locations in the genome: an initial plasmid-encoded dioxygenase and a ring cleavage enzyme and hydrolase encoded on opposite sides of the chromosome.

scholarly journals Novel Pathway for the Degradation of 2-Chloro-4-Nitrobenzoic Acid by Acinetobacter sp. Strain RKJ12

2011 ◽  
Vol 77 (18) ◽  
pp. 6606-6613 ◽  
Author(s):  
Dhan Prakash ◽  
Ravi Kumar ◽  
R. K. Jain ◽  
B. N. Tiwary
Keyword(s):  
Salicylic Acid ◽  
Tricarboxylic Acid Cycle ◽  
Sole Source ◽  
Degradation Pathway ◽  
Ring Cleavage ◽  
Muconic Acid ◽  
Content Type ◽  
Nitrobenzoic Acid ◽  
Catabolic Genes ◽  
Catechol 1,2 Dioxygenase

ABSTRACTThe organismAcinetobactersp. RKJ12 is capable of utilizing 2-chloro-4-nitrobenzoic acid (2C4NBA) as a sole source of carbon, nitrogen, and energy. In the degradation of 2C4NBA by strain RKJ12, various metabolites were isolated and identified by a combination of chromatographic, spectroscopic, and enzymatic activities, revealing a novel assimilation pathway involving both oxidative and reductive catabolic mechanisms. The metabolism of 2C4NBA was initiated by oxidativeorthodehalogenation, leading to the formation of 2-hydroxy-4-nitrobenzoic acid (2H4NBA), which subsequently was metabolized into 2,4-dihydroxybenzoic acid (2,4-DHBA) by a mono-oxygenase with the concomitant release of chloride and nitrite ions. Stoichiometric analysis indicated the consumption of 1 mol O2per conversion of 2C4NBA to 2,4-DHBA, ruling out the possibility of two oxidative reactions. Experiments with labeled H218O and18O2indicated the involvement of mono-oxygenase-catalyzed initial hydrolytic dechlorination and oxidative denitration mechanisms. The further degradation of 2,4-DHBA then proceeds via reductive dehydroxylation involving the formation of salicylic acid. In the lower pathway, the organism transformed salicylic acid into catechol, which was mineralized by theorthoring cleavage catechol-1,2-dioxygenase tocis, cis-muconic acid, ultimately forming tricarboxylic acid cycle intermediates. Furthermore, the studies carried out on a 2C4NBA−derivative and a 2C4NBA+transconjugant demonstrated that the catabolic genes for the 2C4NBA degradation pathway possibly reside on the ∼55-kb transmissible plasmid present in RKJ12.

The metabolism of cyclohexanecarboxylic acid and 3-cyclohexenecarboxylic acid by Pseudomonas putida

1982 ◽  
Vol 28 (12) ◽  
pp. 1324-1329 ◽  
Author(s):  
E. R. Blakley ◽  
B. Papish
Keyword(s):  
Benzoic Acid ◽  
Pseudomonas Putida ◽  
Sole Source ◽  
Oxidation Pathway ◽  
Cyclohexanecarboxylic Acid

A strain of Pseudomonas putida grew rapidly on cyclohexanecarboxylic acid as a sole source of carbon. A CoA-mediated β-oxidation pathway was induced for the metabolism of the compound. The organism could not utilize 3-cyclohexenecarboxylic acid as a sole source of carbon for growth, but cells grown on gluconate in the presence of 3-cyclohexenecarboxylic acid were induced to metabolize cyclohexanecarboxylic acid, benzoic acid, and catechol. Evidence is presented that 3-cyclohexenecarboxylic acid was slowly metabolized by a β-oxidation pathway and by a pathway involving benzoic acid as an intermediate. For this strain of Pseudomonas putida, 3-cyclohexenecarboxylic acid acts as an oxidizable, nongrowth substrate and induces the metabolism of cyclohexanecarboxylic acid and benzoic acid.

Transformation of Alachlor by Microbial Communities

Weed Science ◽  
1990 ◽  
Vol 38 (4-5) ◽  
pp. 416-420 ◽  
Author(s):  
Hone L. Sun ◽  
Thomas J. Sheets ◽  
Frederick T. Corbin
Keyword(s):  
Carbon Source ◽  
Growth Medium ◽  
Basal Medium ◽  
Sole Source ◽  
Ring Cleavage ◽  
Side Chain ◽  
Microbial Culture ◽  
Mixed Microbial Culture ◽  
Alternative Carbon Source ◽  
Thin Layer Chromatographic Analysis

A mixed microbial culture able to transform alachlor at a concentration of 50 μg ml-1was obtained from alachlor-treated soil after an enrichment period of 84 days. The microbial community was composed of seven strains of bacteria. No single isolate was able to utilize alachlor as a sole source of carbon. There was no alachlor left in the enriched culture after a 14-day incubation, but only 12% of the14C-ring-labeled alachlor was converted to14CO2through ring cleavage during 14 days in the basal medium amended with alachlor as a sole carbon source. The presence of sucrose as an alternative carbon source decreased the mineralization potential of the enriched culture, but sucrose increased the mineralizing ability of a three-member mixed culture. Thin-layer chromatographic analysis showed that there were four unidentified metabolites of alachlor produced by the enriched culture. Sucrose decreased the amount of two of the four metabolites. The absence of a noticeable decline in radioactivity beyond the initial 12% suggested that the side chain of alachlor was utilized as carbon source by the enriched culture. Little difference in radioactivity between growth medium and cell-free supernatant of the growth medium suggested that the carbon in the ring was not incorporated into the cells of the degrading microorganisms.

Biochemical pathway and degradation of phthalate ester isomers by bacteria

2005 ◽  
Vol 52 (8) ◽  
pp. 241-248 ◽  
Author(s):  
J.-D. Gu ◽  
J. Li ◽  
Y. Wang
Keyword(s):  
Aromatic Ring ◽  
Enrichment Culture ◽  
Klebsiella Oxytoca ◽  
Sole Source ◽  
Phthalate Ester ◽  
Ring Cleavage ◽  
Mangrove Sediment ◽  
Individual Species ◽  
Dimethyl Phthalate ◽  
Dimethyl Phthalate Ester

Degradation of dimethyl isophthalate (DMI) and dimethyl phthalate ester (DMPE) was investigated using microorganisms isolated from mangrove sediment of Hong Kong Mai Po Nature Reserve. One enrichment culture was capable of utilizing DMI as the sole source of carbon and energy, but none of the bacteria in the enrichment culture was capable of degrading DMI alone. In co-culture of two bacteria, degradation was observed proceeding through monomethyl isophthalate (MMI) ester and isophthalic acid (IPA) before the aromatic ring opening. Using DMI as the sole carbon and energy source, Klebsiella oxytoca Sc and Methylobacterium mesophilicum Sr degraded DMI through the biochemical cooperation. The initial hydrolytic reaction of the ester bond was by K. oxytoca Sc and the next step of transformation was by M. mesophilicum Sr, and IPA was degraded by both of them. In another investigation, a novel bacterium, strain MPsc, was isolated for degradation of dimethyl phthalate ester (DMPE) also from the mangrove sediment. On the basis of phenotypic, biochemical and 16S rDNA gene sequence analyses, the strain MPsc should be considered as a new bacterium at the genus level (8% differences). This strain, together with a Rhodococcus zopfii isolated from the same mangrove sediment, was able to degrade DMPE aerobically. The consortium consisting of the two species degraded 450mg/l DMPE within 3 days as the sole source of carbon and energy, but none of the individual species alone was able to transform DMPE. Furthermore, the biochemical degradation pathway proceeded through monomethyl phthalate (MMP), phthalic acid (PA) and then protocatechuate before aromatic ring cleavage. Our results suggest that degradation of complex organic compounds including DMI and DMPE may be carried out by several members of microorganisms working together in the natural environments.

Degradation of (–)-Ephedrine by Pseudomonas putida Detection of (–)-Ephedrine: NAD+-oxidoreductase from Arthrobacter globiformis

1980 ◽  
Vol 35 (1-2) ◽  
pp. 80-87 ◽  
Author(s):  
E. Klamann ◽  
F. Lingens
Keyword(s):  
Pseudomonas Putida ◽  
Enrichment Culture ◽  
Culture Fluid ◽  
Sole Source ◽  
Culture Technique ◽  
Arthrobacter Globiformis ◽  
Muconic Acid ◽  
Ammonium Sulphate Precipitation ◽  
Wide Range ◽  
Nad Oxidoreductase

Abstract A bacterium utilizing the alkaloid (-)-ephedrine as its sole source of carbon was isolated by an enrichment-culture technique from soil supplemented with 4-benzoyl-1,3-oxazolidinon-(2). The bacterium was identified as Pseudomonasputida by morphological and physiological studies. The following metabolites were isolated from the culture fluid: methylamine, formaldehyde, methyl- benzoylcarbinol (2-hydroxy-1-oxo-1-phenylpropane), benzoid acid, pyrocatechol and cis, cis- muconic acid. A pathway for the degradation of (-)-ephedrine by Pseudomonas putida is proposed and compared with the degradative pathway in Arthrobacter globiformis.The enzyme, which is responsible for the first step in the catabolism of (-)-ephedrine could be demonstrated in extracts from Arthrobacter globiformis. This enzyme catalyses the dehydrogena- tion of (-)-ephedrine yielding phenylacetylcarbinol/methylbenzoylcarbinol and methylamine. It requires NAD+ as cofactor and exhibits optimal activity at pH 11 in 0.1 m glycine/NaOH buffer. The Km value for (-)-ephedrine is 0.02 mM and for NAD+ 0.11 mм, respectively. No remarkable loss of activity is observed following treatment with EDTA. The enzyme has been shown to react with a wide range of ethanolamines. A slight enrichment was obtained by ammonium sulphate precipitation. The name (-)-ephedrine: NAD+-oxidoreductase (deaminating) is proposed.

scholarly journals HipH Catalyzes the Hydroxylation of 4-Hydroxyisophthalate to Protocatechuate in 2,4-Xylenol Catabolism by Pseudomonas putida NCIMB 9866

2015 ◽  
Vol 82 (2) ◽  
pp. 724-731 ◽  
Author(s):  
Hong-Jun Chao ◽  
Yan-Fei Chen ◽  
Ti Fang ◽  
Ying Xu ◽  
Wei E. Huang ◽  
...  
Keyword(s):  
Pseudomonas Putida ◽  
Environmental Protection Agency ◽  
High Performance ◽  
Specific Activity ◽  
Gene Knockout ◽  
Transcriptional Analysis ◽  
Ring Cleavage ◽  
Protection Agency ◽  
Content Type ◽  
Priority Pollutant

ABSTRACTIn addition to growing onp-cresol,Pseudomonas putidaNCIMB 9866 is the only reported strain capable of aerobically growing on 2,4-xylenol, which is listed as a priority pollutant by the U.S. Environmental Protection Agency. Several enzymes involved in the oxidation of thepara-methyl group, as well as the corresponding genes, have previously been reported. The enzyme catalyzing oxidation of the catabolic intermediate 4-hydroxyisophthalate to the ring cleavage substrate protocatechuate was also purified from strain NCIMB 9866, but its genetic determinant is still unavailable. In this study, the genehipH, encoding 4-hydroxyisophthalate hydroxylase, from strain NCIMB 9866 was cloned by transposon mutagenesis. Purified recombinant HipH-His6was found to be a dimer protein with a molecular mass of approximately 110 kDa. HipH-His6catalyzed the hydroxylation of 4-hydroxyisophthalate to protocatechuate with a specific activity of 1.54 U mg−1and showed apparentKmvalues of 11.40 ± 3.05 μM for 4-hydroxyisophthalate with NADPH and 11.23 ± 2.43 μM with NADH and similarKmvalues for NADPH and NADH (64.31 ± 13.16 and 72.76 ± 12.06 μM, respectively). The identity of protocatechuate generated from 4-hydroxyisophthalate hydroxylation by HipH-His6has also been confirmed by high-performance liquid chromatography and mass spectrometry. Gene transcriptional analysis, gene knockout, and complementation indicated thathipHis essential for 2,4-xylenol catabolism but not forp-cresol catabolism in this strain. This fills a gap in our understanding of the gene that encodes a critical step in 2,4-xylenol catabolism and also provides another example of biochemical and genetic diversity of microbial catabolism of structurally similar compounds.

scholarly journals Bacterial Degradation of N,N-Diethyl-m-Toluamide (DEET): Cloning and Heterologous Expression of DEET Hydrolase

2007 ◽  
Vol 73 (9) ◽  
pp. 3105-3108 ◽  
Author(s):  
Giomar Rivera-Cancel ◽  
Daniela Bocioaga ◽  
Anthony G. Hay
Keyword(s):  
Carbon Source ◽  
Heterologous Expression ◽  
Pseudomonas Putida ◽  
Sole Carbon Source ◽  
Bacterial Degradation ◽  
Ring Cleavage

ABSTRACT Pseudomonas putida DTB grew aerobically with N,N-diethyl-m-toluamide (DEET) as a sole carbon source, initially breaking it down into 3-methylbenzoate and diethylamine. The former was further metabolized via 3-methylcatechol and meta ring cleavage. A gene from DTB, dthA, was heterologously expressed and shown to encode the ability to hydrolyze DEET into 3-methylbenzoate and diethylamine.

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