Diverse Reductive Dehalogenases Are Associated with Clostridiales-Enriched Microcosms Dechlorinating 1,2-Dichloroethane

The achievement of successful biostimulation of active microbiomes for the cleanup of a polluted site is strictly dependent on the knowledge of the key microorganisms equipped with the relevant catabolic genes responsible for the degradation process. In this work, we present the characterization of the bacterial community developed in anaerobic microcosms after biostimulation with the electron donor lactate of groundwater polluted with 1,2-dichloroethane (1,2-DCA). Through a multilevel analysis, we have assessed (i) the structural analysis of the bacterial community; (ii) the identification of putative dehalorespiring bacteria; (iii) the characterization of functional genes encoding for putative 1,2-DCA reductive dehalogenases (RDs). Following the biostimulation treatment, the structure of the bacterial community underwent a notable change of the main phylotypes, with the enrichment of representatives of the orderClostridiales. Through PCR targeting conserved regions within known RD genes, four novel variants of RDs previously associated with the reductive dechlorination of 1,2-DCA were identified in the metagenome of the Clostridiales-dominated bacterial community.

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Bacterial Catabolism of β-Hydroxypropiovanillone and β-Hydroxypropiosyringone Produced in the Reductive Cleavage of Arylglycerol-β-Aryl Ether in Lignin

Applied and Environmental Microbiology ◽

10.1128/aem.02670-17 ◽

2018 ◽

Vol 84 (7) ◽

Cited By ~ 4

Author(s):

Yudai Higuchi ◽

Shogo Aoki ◽

Hiroki Takenami ◽

Naofumi Kamimura ◽

Kenji Takahashi ◽

...

Keyword(s):

Value Added ◽

Lignin Biodegradation ◽

Gene Products ◽

Aryl Ether ◽

Ether Linkage ◽

Content Type ◽

Catabolic Genes ◽

Genes Encoding ◽

Stereospecific Reaction

ABSTRACTSphingobiumsp. strain SYK-6 converts four stereoisomers of arylglycerol-β-guaiacyl ether into achiral β-hydroxypropiovanillone (HPV) via three stereospecific reaction steps. Here, we determined the HPV catabolic pathway and characterized the HPV catabolic genes involved in the first two steps of the pathway. In SYK-6 cells, HPV was oxidized to vanilloyl acetic acid (VAA) via vanilloyl acetaldehyde (VAL). The resulting VAA was further converted into vanillate through the activation of VAA by coenzyme A. A syringyl-type HPV analog, β-hydroxypropiosyringone (HPS), was also catabolized via the same pathway. SLG_12830 (hpvZ), which belongs to the glucose-methanol-choline oxidoreductase family, was isolated as the HPV-converting enzyme gene. AnhpvZmutant completely lost the ability to convert HPV and HPS, indicating thathpvZis essential for the conversion of both the substrates. HpvZ produced inEscherichia colioxidized both HPV and HPS and other 3-phenyl-1-propanol derivatives. HpvZ localized to both the cytoplasm and membrane of SYK-6 and used ubiquinone derivatives as electron acceptors. Thirteen gene products of the 23 aldehyde dehydrogenase (ALDH) genes in SYK-6 were able to oxidize VAL into VAA. Mutant analyses suggested that multiple ALDH genes, including SLG_20400, contribute to the conversion of VAL. We examined whether the genes encoding feruloyl-CoA synthetase (ferA) and feruloyl-CoA hydratase/lyase (ferBandferB2) are involved in the conversion of VAA. Only FerA exhibited activity toward VAA; however, disruption offerAdid not affect VAA conversion. These results indicate that another enzyme system is involved in VAA conversion.IMPORTANCECleavage of the β-aryl ether linkage is the most essential process in lignin biodegradation. Although the bacterial β-aryl ether cleavage pathway and catabolic genes have been well documented, there have been no reports regarding the catabolism of HPV or HPS, the products of cleavage of β-aryl ether compounds. HPV and HPS have also been found to be obtained from lignin by chemoselective catalytic oxidation by 2,3-dichloro-5,6-dicyano-1,4-benzoquinone/tert-butyl nitrite/O2, followed by cleavage of the β-aryl ether with zinc. Therefore, value-added chemicals are expected to be produced from these compounds. In this study, we determined the SYK-6 catabolic pathways for HPV and HPS and identified the catabolic genes involved in the first two steps of the pathways. Since SYK-6 catabolizes HPV through 2-pyrone-4,6-dicarboxylate, which is a building block for functional polymers, characterization of HPV catabolism is important not only for understanding the bacterial lignin catabolic system but also for lignin utilization.

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Recent Advanced Technologies for the Characterization of Xenobiotic-Degrading Microorganisms and Microbial Communities

Frontiers in Bioengineering and Biotechnology ◽

10.3389/fbioe.2021.632059 ◽

2021 ◽

Vol 9 ◽

Author(s):

Sandhya Mishra ◽

Ziqiu Lin ◽

Shimei Pang ◽

Wenping Zhang ◽

Pankaj Bhatt ◽

...

Keyword(s):

Microbial Communities ◽

Complex Mixture ◽

Nitrogen Sources ◽

Degradation Process ◽

Microbial Populations ◽

Novel Proteins ◽

Catabolic Genes ◽

Xenobiotic Compounds ◽

Metabolic Activities

Global environmental contamination with a complex mixture of xenobiotics has become a major environmental issue worldwide. Many xenobiotic compounds severely impact the environment due to their high toxicity, prolonged persistence, and limited biodegradability. Microbial-assisted degradation of xenobiotic compounds is considered to be the most effective and beneficial approach. Microorganisms have remarkable catabolic potential, with genes, enzymes, and degradation pathways implicated in the process of biodegradation. A number of microbes, including Alcaligenes, Cellulosimicrobium, Microbacterium, Micrococcus, Methanospirillum, Aeromonas, Sphingobium, Flavobacterium, Rhodococcus, Aspergillus, Penecillium, Trichoderma, Streptomyces, Rhodotorula, Candida, and Aureobasidium, have been isolated and characterized, and have shown exceptional biodegradation potential for a variety of xenobiotic contaminants from soil/water environments. Microorganisms potentially utilize xenobiotic contaminants as carbon or nitrogen sources to sustain their growth and metabolic activities. Diverse microbial populations survive in harsh contaminated environments, exhibiting a significant biodegradation potential to degrade and transform pollutants. However, the study of such microbial populations requires a more advanced and multifaceted approach. Currently, multiple advanced approaches, including metagenomics, proteomics, transcriptomics, and metabolomics, are successfully employed for the characterization of pollutant-degrading microorganisms, their metabolic machinery, novel proteins, and catabolic genes involved in the degradation process. These technologies are highly sophisticated, and efficient for obtaining information about the genetic diversity and community structures of microorganisms. Advanced molecular technologies used for the characterization of complex microbial communities give an in-depth understanding of their structural and functional aspects, and help to resolve issues related to the biodegradation potential of microorganisms. This review article discusses the biodegradation potential of microorganisms and provides insights into recent advances and omics approaches employed for the specific characterization of xenobiotic-degrading microorganisms from contaminated environments.

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Characterization of an L-Ascorbate Catabolic Pathway with Unprecedented Enzymatic Transformations

10.26434/chemrxiv.9808241.v1 ◽

2019 ◽

Author(s):

Tyler Stack ◽

Katelyn Morrison ◽

Thomas Dettmer ◽

Brendan Wille ◽

Chan Kim ◽

...

Keyword(s):

Ralstonia Eutropha ◽

Sequence Similarity ◽

Catabolic Genes ◽

Genes Encoding ◽

Benzilic Acid ◽

Amidohydrolase Superfamily ◽

Sequence Similarity Networks

L-Ascorbate (vitamin C) is ubiquitous in both our diet and the environment. Ralstonia eutropha H16 (Cupriavidus necator ATCC 17699) uses L-ascorbate as sole carbon source but lacks the genes encoding the known catabolic pathways. RNAseq identified eight candidate catabolic genes. Sequence similarity networks and genome neighborhood networks guided predictions for function of the encoded proteins; the predictions were confirmed by in vitro assays and in vivo growth phenotypes of gene deletion mutants. L-Ascorbate, a lactone, is oxidized and ring-opened by enzymes in the cytochrome b561 and gluconolactonase families, respectively, to form 2,3-diketo-L-gulonate. A protein predicted to have a WD40-like fold catalyzes an unprecedented benzilic acid rearrangement involving migration of a carboxylate group to form 2-carboxy-L-lyxonolactone; the lactone is hydrolyzed by a member of the amidohydrolase superfamily to yield 2-carboxy-L-lyxonate. A member of the PdxA family of oxidative decarboxylases catalyzes a novel decarboxylation that uses NAD+ catalytically. The product, L-lyxonate, is catabolized to alpha-ketoglutarate by a previously characterized pathway.

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Molecular Characterization of New Haplotype of Genus Sarcocystis in Seabirds from Magdalena Island, Southern Chile

Animals ◽

10.3390/ani11020245 ◽

2021 ◽

Vol 11 (2) ◽

pp. 245

Author(s):

Igor C. L. Acosta ◽

Solange M. Gennari ◽

Horwald A. B. Llano ◽

Sebastián Muñoz-Leal ◽

Rodrigo M. Soares

Keyword(s):

Nested Pcr ◽

Cytochrome Oxidase Subunit ◽

Gene Encoding ◽

Marine Fauna ◽

Spheniscus Magellanicus ◽

Larus Dominicanus ◽

B1 Gene ◽

Genes Encoding ◽

Pcr Targeting

Evidence of sarcocystid infection was investigated in samples of 16 penguins (Spheniscus. magellanicus), four Dominican gulls (Larus dominicanus) and two Chilean skuas (Stercorarius chilensis) found in Madalenas Islands, Chile, in 2017. Samples of skeletal muscle, cardiac muscle and brain from all birds were screened by a pan-sarcocystid nested-PCR targeting a short fragment of the gene encoding the small ribosomal unit (nPCR-18Sa). The only two positive samples by nPCR-18Sa, both from skuas, were tested by a nested-PCR directed to the internal transcribed spacer 1 (nPCR-ITS1), also a pan-sarcocystidae nested-PCR, and to a nested-PCR directed to the B1 gene (nPCR-B1), for the exclusive detection of Toxoplasma gondii. The two nPCR-18Sa-positive samples were nPCR-ITS1-positive and nPCR-B1-negative. The nPCR-ITS1 nucleotide sequences from the two skuas, which were identical to each other, were revealed closely related to homologous sequences of Sarcocystis halieti, species found in seabirds of northern hemisphere. Larger fragments of genes encoding 18S and partial sequences of genes coding for cytochrome oxidase subunit 1 were also analyzed, corroborating ITS1 data. The haplotypes found in the skuas are unprecedent and closely related to species that use birds as the definitive host. Further studies need to be carried out to detect, identify and isolate this parasite to understand the epidemiology of the infection and its impact on the health of marine fauna.

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Isolation and characterization of two specific regulatory Aspergillus niger mutants shows antagonistic regulation of arabinan and xylan metabolism

Microbiology ◽

10.1099/mic.0.25993-0 ◽

2003 ◽

Vol 149 (5) ◽

pp. 1183-1191 ◽

Cited By ~ 39

Author(s):

Marco J. L. de Groot ◽

Peter J. I. van de Vondervoort ◽

Ronald P. de Vries ◽

Patricia A. vanKuyk ◽

George J. G. Ruijter ◽

...

Keyword(s):

Aspergillus Niger ◽

Regulatory System ◽

Gene Encoding ◽

Isolation And Characterization ◽

Catabolic Genes ◽

Genes Encoding ◽

Intracellular Activities ◽

Xylulose Kinase ◽

Encoding Genes

This paper describes two Aspergillus niger mutants (araA and araB) specifically disturbed in the regulation of the arabinanase system in response to the presence of l-arabinose. Expression of the three known l-arabinose-induced arabinanolytic genes, abfA, abfB and abnA, was substantially decreased or absent in the araA and araB strains compared to the wild-type when incubated in the presence of l-arabinose or l-arabitol. In addition, the intracellular activities of l-arabitol dehydrogenase and l-arabinose reductase, involved in l-arabinose catabolism, were decreased in the araA and araB strains. Finally, the data show that the gene encoding d-xylulose kinase, xkiA, is also under control of the arabinanolytic regulatory system. l-Arabitol, most likely the true inducer of the arabinanolytic and l-arabinose catabolic genes, accumulated to a high intracellular concentration in the araA and araB mutants. This indicates that the decrease of expression of the arabinanolytic genes was not due to lack of inducer accumulation. Therefore, it is proposed that the araA and araB mutations are localized in positive-acting components of the regulatory system involved in the expression of the arabinanase-encoding genes and the genes encoding the l-arabinose catabolic pathway.

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Characterization of an L-Ascorbate Catabolic Pathway with Unprecedented Enzymatic Transformations

10.26434/chemrxiv.9808241 ◽

2019 ◽

Author(s):

Tyler Stack ◽

Katelyn Morrison ◽

Thomas Dettmer ◽

Brendan Wille ◽

Chan Kim ◽

...

Keyword(s):

Ralstonia Eutropha ◽

Sequence Similarity ◽

Catabolic Genes ◽

Genes Encoding ◽

Benzilic Acid ◽

Amidohydrolase Superfamily ◽

Sequence Similarity Networks

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l-Rhamnose Transport Is Sugar Kinase (RhaK) Dependent in Rhizobium leguminosarum bv. trifolii

Journal of Bacteriology ◽

10.1128/jb.01032-07 ◽

2007 ◽

Vol 189 (23) ◽

pp. 8437-8446 ◽

Cited By ~ 9

Author(s):

Jason S. Richardson ◽

Ivan J. Oresnik

Keyword(s):

Abc Transporter ◽

Rhizobium Leguminosarum ◽

Nodule Occupancy ◽

Vitro Assay ◽

Catabolic Genes ◽

Genes Encoding ◽

Pentose Sugar ◽

Sugar Kinase

ABSTRACT Strains of Rhizobium leguminosarum which are unable to catabolize l-rhamnose, a methyl-pentose sugar, are compromised in the ability to compete for nodule occupancy versus wild-type strains. Previous characterization of the 11-kb region necessary for the utilization of rhamnose identified a locus carrying catabolic genes and genes encoding the components of an ABC transporter. Genetic evidence suggested that the putative kinase RhaK carried out the first step in the catabolism of rhamnose. Characterization of this kinase led to the observation that strains carrying rhamnose kinase mutations were unable to transport rhamnose into the cell. The absence of a functional rhamnose kinase did not stop the transcription and translation of the ABC transporter components. By developing an in vitro assay for RhaK activity, we have been able to show that (i) RhaK activity is consistent with RhaK phosphorylating rhamnose and (ii) biochemical activity of RhaK is necessary for rhamnose transport.

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Approaches to assess the biodiversity of bacteria in natural habitats

Biochemical Society Transactions ◽

10.1042/bst0340169 ◽

2006 ◽

Vol 34 (1) ◽

pp. 169-173 ◽

Cited By ~ 3

Author(s):

C. Rösch ◽

S. Eilmus ◽

H. Bothe

Keyword(s):

Bacterial Community ◽

Environmental Samples ◽

Target Genes ◽

Pcr Amplification ◽

Restriction Enzymes ◽

Functional Genes ◽

Natural Habitats ◽

Uncultured Bacteria ◽

Computer Based

Any attempt to characterize a bacterial community and their functional genes coding for enzymes of the nitrogen cycle is faced with its extreme biodiversity. Novel techniques, based on PCR amplification of target genes in DNA from environmental samples, have been developed for characterizing both cultured and as yet uncultured bacteria in the last few years. Computer-based assignment tools have now been developed utilizing terminal restriction fragments obtained from digestions with multiple restriction enzymes. Such programs allow the gross characterization of bacterial life in any complex bacterial community with confidence.

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