Identification of Camphor Oxidation and Reduction Products in Pseudomonas putida: New Activity of the Cytochrome P450cam System

Prasad, Brinda; Rojubally, Adina; Plettner, Erika

doi:10.1007/s10886-011-9959-7

Cited by 13 publications

(18 citation statements)

References 31 publications

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“…In vivo , this occurs when cultures are poorly aerated [16] and, in vitro , this occurs when the buffer is sparged with argon in an open vial. In contrast, the known oxidation products 10 and 11 form at high O 2 concentrations (∼9 mg/L = 284 µM).…”

Section: Resultsmentioning

confidence: 99%

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Water Oxidation by a Cytochrome P450: Mechanism and Function of the Reaction

et al. 2013

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P450cam (CYP101A1) is a bacterial monooxygenase that is known to catalyze the oxidation of camphor, the first committed step in camphor degradation, with simultaneous reduction of oxygen (O2). We report that P450cam catalysis is controlled by oxygen levels: at high O2 concentration, P450cam catalyzes the known oxidation reaction, whereas at low O2 concentration the enzyme catalyzes the reduction of camphor to borneol. We confirmed, using 17O and 2H NMR, that the hydrogen atom added to camphor comes from water, which is oxidized to hydrogen peroxide (H2O2). This is the first time a cytochrome P450 has been observed to catalyze oxidation of water to H2O2, a difficult reaction to catalyze due to its high barrier. The reduction of camphor and simultaneous oxidation of water are likely catalyzed by the iron-oxo intermediate of P450cam, and we present a plausible mechanism that accounts for the 1∶1 borneol:H2O2 stoichiometry we observed. This reaction has an adaptive value to bacteria that express this camphor catabolism pathway, which requires O2, for two reasons: 1) the borneol and H2O2 mixture generated is toxic to other bacteria and 2) borneol down-regulates the expression of P450cam and its electron transfer partners. Since the reaction described here only occurs under low O2 conditions, the down-regulation only occurs when O2 is scarce.

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Section: Resultsmentioning

confidence: 99%

“…1b) [16]. We have interpreted this reaction to give P. putida an ecological advantage over other non-camphor metabolising bacteria because borneol is bactericidal to non-P450 containing bacteria, but not to P. putida [16].…”

Section: Introductionmentioning

confidence: 98%

Water Oxidation by a Cytochrome P450: Mechanism and Function of the Reaction

et al. 2013

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“…aldo-keto-reductases, carbonyl-reductases) [49,50], but focussed studies with molecules comparable to oxo-1,8-cineoles have, as of yet, not been accomplished. However, some bacterial enzymes have been reported to be responsible for very similar reactions such as the reduction of camphor to borneol [51], and human carbonyl reductases have such a broad substrate specificity [52] that it is well conceivable that such a back-reaction of oxo- to hydroxy- 1,8-cineole might take place, as also proposed by Ishida et al for thujone and carvone [53]. …”

Section: Discussionmentioning

confidence: 99%

Characterisation of the Metabolites of 1,8-Cineole Transferred into Human Milk: Concentrations and Ratio of Enantiomers

Kirsch

Buettner

2013

Metabolites

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1,8-Cineole is a widely distributed odorant that also shows physiological effects, but whose human metabolism has hitherto not been extensively investigated. The aim of the present study was, thus, to characterise the metabolites of 1,8-cineole, identified previously in human milk, after the oral intake of 100 mg of this substance. Special emphasis was placed on the enantiomeric composition of the metabolites since these data may provide important insights into potential biotransformation pathways, as well as potential biological activities of these substances, for example on the breastfed child. The volatile fraction of the human milk samples was therefore isolated via Solvent Assisted Flavour Evaporation (SAFE) and subjected to gas chromatography-mass spectrometry (GC-MS). The absolute concentrations of each metabolite were determined by matrix calibration with an internal standard, and the ratios of enantiomers were analysed on chiral capillaries. The concentrations varied over a broad range, from traces in the upper ng/kg region up to 40 µg/kg milk, with the exception of the main metabolite α2-hydroxy-1,8-cineole that showed concentrations of 100–250 µg/kg. Also, large inter- and intra-individual variations were recorded for the enantiomers, with nearly enantiomerically pure α2-hydroxy- and 3-oxo-1,8-cineole, while all other metabolites showed ratios of ~30:70 to 80:20.

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“…CBMAI 1197 is an efficient biocatalyst that promotes mono-biohydroxylation of rac-camphor (1). The main biotransformation product was 6-endo-hydroxycamphor (3) reaching 49% after 5 days of reaction followed by 5-exohydroxycamphor (5), 5-endo-hydroxycamphor (6), 6-exohydroxycamphor (4) and 8-hydroxycamphor (7). Chemical modifications in non-activated carbon atoms such as sp 3 camphor carbons usually cannot be done by conventional organic synthesis.…”

Section: Discussionmentioning

confidence: 99%

“…For example, cytochrome P450 from Pseudomonas putida catalyzed the selective oxidation of camphor to 5-exo-hydroxycamphor. 7 There are few articles in the literature describing monohydroxylation of camphor. Nakahashi and Miyazawa 8 used Salmonella typhimurium OY1000/2A6 of human cytochrome P450, employing the cofactor NADPH-P450 reductase.…”

Section: Introductionmentioning

confidence: 99%

Multiple Monohydroxylation Products from rac-Camphor by Marine Fungus Botryosphaeria sp. Isolated from Marine Alga Bostrychia radicans

Jesus

Jeller

Debonsi

et al. 2016

J. Braz. Chem. Soc.

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This manuscript describes the biooxidation of rac-camphor using whole cells of marine-derived fungus Botryosphaeria sp. CBMAI 1197. The main biotransformation products of this monoterpene were achieved via a hydroxylation reaction and occurred with 5 days of rac-camphor incubation. Products were identified by means of gas chromatography mass spectrometry (GC-MS) and nuclear magnetic resonance (NMR) data. The major hydroxylated products were 6-endo-hydroxycamphor, 6-exo-hydroxycamphor, 5-exo-hydroxycamphor, 5-endo-hydroxycamphor, 3-exo-hydroxycamphor and 8-hydroxycamphor. The 6-exo-hydroxycamphor was obtained through a retro-aldol reaction when 6-endo-hydroxycamphor was maintained in presence of CDCl 3 ; this isomerization was confirmed by 1 H NMR and GC-MS data.

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Identification of Camphor Oxidation and Reduction Products in Pseudomonas putida: New Activity of the Cytochrome P450cam System

Cited by 13 publications

References 31 publications

Water Oxidation by a Cytochrome P450: Mechanism and Function of the Reaction

Water Oxidation by a Cytochrome P450: Mechanism and Function of the Reaction

Characterisation of the Metabolites of 1,8-Cineole Transferred into Human Milk: Concentrations and Ratio of Enantiomers

Multiple Monohydroxylation Products from rac-Camphor by Marine Fungus Botryosphaeria sp. Isolated from Marine Alga Bostrychia radicans

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