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Metabolic pathway involved in 2-methyl-6-ethylaniline degradation by Sphingobium sp. strain MEA3-1 and cloning of the novel flavin-dependent monooxygenase system meaBA.
- Source :
-
Applied and environmental microbiology [Appl Environ Microbiol] 2015 Dec; Vol. 81 (24), pp. 8254-64. Date of Electronic Publication: 2015 Sep 18. - Publication Year :
- 2015
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Abstract
- 2-Methyl-6-ethylaniline (MEA) is the main microbial degradation intermediate of the chloroacetanilide herbicides acetochlor and metolachlor. Sphingobium sp. strain MEA3-1 can utilize MEA and various alkyl-substituted aniline and phenol compounds as sole carbon and energy sources for growth. We isolated the mutant strain MEA3-1Mut, which converts MEA only to 2-methyl-6-ethyl-hydroquinone (MEHQ) and 2-methyl-6-ethyl-benzoquinone (MEBQ). MEA may be oxidized by the P450 monooxygenase system to 4-hydroxy-2-methyl-6-ethylaniline (4-OH-MEA), which can be hydrolytically spontaneously deaminated to MEBQ or MEHQ. The MEA microbial metabolic pathway was reconstituted based on the substrate spectra and identification of the intermediate metabolites in both the wild-type and mutant strains. Plasmidome sequencing indicated that both strains harbored 7 plasmids with sizes ranging from 6,108 bp to 287,745 bp. Among the 7 plasmids, 6 were identical, and pMEA02' in strain MEA3-1Mut lost a 37,000-bp fragment compared to pMEA02 in strain MEA3-1. Two-dimensional electrophoresis (2-DE) and protein mass fingerprinting (PMF) showed that MEA3-1Mut lost the two-component flavin-dependent monooxygenase (TC-FDM) MeaBA, which was encoded by a gene in the lost fragment of pMEA02. MeaA shared 22% to 25% amino acid sequence identity with oxygenase components of some TC-FDMs, whereas MeaB showed no sequence identity with the reductase components of those TC-FDMs. Complementation with meaBA in MEA3-1Mut and heterologous expression in Pseudomonas putida strain KT2440 resulted in the production of an active MEHQ monooxygenase.<br /> (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)
- Subjects :
- Acetamides metabolism
Amino Acid Sequence
Base Sequence
Biodegradation, Environmental
DNA, Bacterial genetics
Delftia enzymology
Delftia genetics
Electrophoresis, Gel, Two-Dimensional
Metabolic Networks and Pathways genetics
Molecular Sequence Data
Oxygenases metabolism
Peptide Mapping
Pseudomonas putida metabolism
Sequence Analysis, DNA
Sphingomonadaceae enzymology
Sphingomonadaceae genetics
Delftia metabolism
Oxygenases genetics
Sphingomonadaceae metabolism
Toluidines metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 1098-5336
- Volume :
- 81
- Issue :
- 24
- Database :
- MEDLINE
- Journal :
- Applied and environmental microbiology
- Publication Type :
- Academic Journal
- Accession number :
- 26386060
- Full Text :
- https://doi.org/10.1128/AEM.01883-15