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1.
Proc Natl Acad Sci U S A ; 117(32): 19228-19236, 2020 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-32703810

RESUMO

The ATP-binding cassette (ABC) transporter of mitochondria (Atm1) mediates iron homeostasis in eukaryotes, while the prokaryotic homolog from Novosphingobium aromaticivorans (NaAtm1) can export glutathione derivatives and confer protection against heavy-metal toxicity. To establish the structural framework underlying the NaAtm1 transport mechanism, we determined eight structures by X-ray crystallography and single-particle cryo-electron microscopy in distinct conformational states, stabilized by individual disulfide crosslinks and nucleotides. As NaAtm1 progresses through the transport cycle, conformational changes in transmembrane helix 6 (TM6) alter the glutathione-binding site and the associated substrate-binding cavity. Significantly, kinking of TM6 in the post-ATP hydrolysis state stabilized by MgADPVO4 eliminates this cavity, precluding uptake of glutathione derivatives. The presence of this cavity during the transition from the inward-facing to outward-facing conformational states, and its absence in the reverse direction, thereby provide an elegant and conceptually simple mechanism for enforcing the export directionality of transport by NaAtm1. One of the disulfide crosslinked NaAtm1 variants characterized in this work retains significant glutathione transport activity, suggesting that ATP hydrolysis and substrate transport by Atm1 may involve a limited set of conformational states with minimal separation of the nucleotide-binding domains in the inward-facing conformation.


Assuntos
Transportadores de Cassetes de Ligação de ATP/química , Proteínas de Bactérias/química , Sphingomonadaceae/metabolismo , Transportadores de Cassetes de Ligação de ATP/genética , Transportadores de Cassetes de Ligação de ATP/metabolismo , Trifosfato de Adenosina/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Sítios de Ligação , Cristalografia por Raios X , Glutationa/química , Glutationa/metabolismo , Ferro/metabolismo , Domínios Proteicos , Sphingomonadaceae/química , Sphingomonadaceae/genética
2.
Chemosphere ; 258: 127322, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-32563915

RESUMO

The dissipation of racemic, R-, and S- dichlorprop (DCPP) in four soils were studied in the laboratory. The half-lives of racemic DCPP were from 10.5 to 19.8 days. Preferential degradation of R- or S-DCPP was detected in all soils, even in one soil that the apparent enantiomeric fraction remained constant during incubation. The enantiomerization of DCPP was found to proceed in both directions, except in forest soil that no enantiomerization of S- to R-DCPP was observed. The isomerization equilibrium constant (K = kRS/kSR) in two vegetable soils were 0.54 and 0.53, respectively, favoring herbicidally active R enantiomer, while in paddy soil K was 1.60, favoring an inversion of R into S enantiomer. Real-time PCR showed that the rdpA gene was not detected in all indigenous and DCPP amended microcosms probably because of relative short incubation time and low amendment concentrations. In contrast, the sdpA gene was present in indigenous soils and significantly elevated after DCPP addition with the highest relative abundance around day 10 in all microcosms. Illumina sequencing of the 16S rRNA gene showed that the relative abundance of Proteobacteria significantly increased in all DCPP treated soils. DCPP-degrading related families, Sphingomonadaceae and Comamonadaceae, enhanced in all soils, while Burkholderiaceae elevated only in paddy soil with preferential degradation of S-DCPP and Pseudomonadaceae only in forest soil with R-enantiomer preference. The sdpA gene sequencing revealed that about 92%-99% of bacteria harboring sdpA genes in studied soils belong to Alphaproteobacteria.


Assuntos
Ácido 2,4-Diclorofenoxiacético/análogos & derivados , Biodegradação Ambiental , Poluentes do Solo/metabolismo , Ácido 2,4-Diclorofenoxiacético/metabolismo , Comamonadaceae/metabolismo , Genes Bacterianos , Herbicidas/análise , RNA Ribossômico 16S , Solo , Microbiologia do Solo , Poluentes do Solo/análise , Sphingomonadaceae/metabolismo , Estereoisomerismo
3.
Chemosphere ; 255: 126906, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32387905

RESUMO

Enzymatic function of MlrC from Novosphingobium sp. THN1 (i.e., THN1-MlrC) towards linearized microcystins (MCs) and structural/physic-chemical properties of MlrC enzyme deserved urgent research, and heterologous expression (HE) optimization for MlrC is yet to be solved. This study achieved HE of THN1-MlrC by rapid-efficiently constructing HE system, and revealed that THN1-MlrC can degrade linearized MC-LR and linearized MC-RR to produce Adda, providing direct evidence for catalytic function of THN-MlrC and its ecological implication in MC-detoxification. Consequently, to maximize THN1-MlrC expression for production and application, induction conditions for HE of THN1-MlrC was optimized as at 0.1 mM of isopropyl-ß-D-thiogalactoside and 30 °C for 8 h, which could be widely applicable for heterologous production of other MlrC homologs. Using bioinformatics and site-mutation experiment, THN1-MlrC was evaluated as a cytoplasm-locating hydrophilic protein with theoretical isoelectric point of 5.57, and contained six verified active sites Glu39, His133, Asp167, His169, His191 and Asp332 in two domains of its 3D structure, among which Glu39, His133 and Asp332 were newly-discovered ones here. The Glu39, His133, Asp167, His169 and His191 gathered more closely in 3D structure than in amino acid sequence, while they and Asp332 surrounded protein center to constitute a potential active pocket for mediating linearized MCs degradation. Due to MlrC sequence homology and conservative active sites, structural/physic-chemical characteristics of THN1-MlrC presented in this study provided a helpful reference for other MlrC homologs of diverse bacteria. This study shed novel insights for functional-structural relationships of THN1-MlrC during MC-biodegradation, and was crucial for research and practical applications in MC-decontamination.


Assuntos
Biodegradação Ambiental , Microcistinas/metabolismo , Biologia Computacional , Mutagênese Sítio-Dirigida , Sphingomonadaceae/metabolismo
4.
Appl Environ Microbiol ; 86(12)2020 06 02.
Artigo em Inglês | MEDLINE | ID: mdl-32303545

RESUMO

Carboxylesterase PytH, isolated from the pyrethroid-degrading bacterium Sphingobium faniae JZ-2, could rapidly hydrolyze the ester bond of a wide range of pyrethroid pesticides, including permethrin, fenpropathrin, cypermethrin, fenvalerate, deltamethrin, cyhalothrin, and bifenthrin. To elucidate the catalytic mechanism of PytH, we report here the crystal structures of PytH with bifenthrin (BIF) and phenylmethylsulfonyl fluoride (PMSF) and two PytH mutants. Though PytH shares low sequence identity with reported α/ß-hydrolase fold proteins, the typical triad catalytic center with Ser-His-Asp triad (Ser78, His230, and Asp202) is present and vital for the hydrolase activity. However, no contact was found between Ser78 and His230 in the structures we solved, which may be due to the fact that the PytH structures we determined are in their inactive or low-activity forms. The structure of PytH is composed of a core domain and a lid domain; some hydrophobic amino acid residues surrounding the substrate from both domains form a deeper and wider hydrophobic pocket than its homologous structures. This indicates that the larger hydrophobic pocket makes PytH fit for its larger substrate binding; both lid and core domains are involved in substrate binding, and the lid domain-induced core domain movement may make the active center correctly positioned with substrates.IMPORTANCE Pyrethroid pesticides are widely applied in agriculture and household; however, extensive use of these pesticides also causes serious environmental and health problems. The hydrolysis of pyrethroids by carboxylesterases is the major pathway of microbial degradation of pyrethroids, but the structure of carboxylesterases and its catalytic mechanism are still unknown. Carboxylesterase PytH from Sphingobium faniae JZ-2 could effectively hydrolyze a wide range of pyrethroid pesticides. The crystal structures of PytH are solved in this study. This showed that PytH belongs to the α/ß-hydrolase fold proteins with typical catalytic Ser-His-Asp triad, though PytH has a low sequence identity (about 20%) with them. The special large hydrophobic binding pocket enabled PytH to bind bigger pyrethroid family substrates. Our structures shed light on the substrate selectivity and the future application of PytH and deepen our understanding of α/ß-hydrolase members.


Assuntos
Proteínas de Bactérias/genética , Hidrolases de Éster Carboxílico/genética , Inseticidas/metabolismo , Fluoreto de Fenilmetilsulfonil/metabolismo , Piretrinas/metabolismo , Sphingomonadaceae/genética , Proteínas de Bactérias/metabolismo , Hidrolases de Éster Carboxílico/metabolismo , Análise de Sequência de DNA , Sphingomonadaceae/metabolismo
5.
J Biosci Bioeng ; 130(1): 71-75, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32238321

RESUMO

Lignosulfonate is a by-product of the cooking process by sulfite pulping for paper manufacturing. The treatment of wood chips by various salts of sulfurous acid solubilizes lignin to produce a cellulose-rich wood pulp. Developing a technique for the conversion of lignosulfonate by-product to high value materials has an important industrial utility. Sphingobium sp. strain SYK-6, which was isolated from pulping wastewater, is one of the best enzymatically or genetically characterized bacteria for degrading lignin-derived aromatics. We have previously established a system for the production of 2-pyrone-4,6-dicarboxylic acid (PDC), a novel platform chemical that can produce a variety of bio-based polymers, by introducing of ligA, ligB, and ligC genes from SYK-6 into a mutant strain of Pseudomonas putida PpY1100. In this study, extracts from lignosulfonates, which were desulphonated and depolymerized by alkaline oxidation, were evaluated as substrates for microbiological conversion to PDC by the transgenic bacteria.


Assuntos
Lignina/metabolismo , Extratos Vegetais/metabolismo , Pseudomonas putida/metabolismo , Pironas/metabolismo , Sphingomonadaceae/metabolismo , Celulose/metabolismo , Ácidos Dicarboxílicos/metabolismo , Pseudomonas putida/genética , Sphingomonadaceae/genética , Resíduos/análise
6.
Artif Cells Nanomed Biotechnol ; 48(1): 672-682, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32075448

RESUMO

The present study highlights the biological synthesis of silver nanoparticles (AgNPs) using Sphingobium sp. MAH-11 and also their antibacterial mechanisms against drug-resistant pathogenic microorganisms. The nanoparticle synthesis method used in this study was reliable, facile, rapid, cost-effective and ecofriendly. The AgNPs exhibited the highest absorbance at 423 nm. The TEM image expressed spherical shape of AgNPs and the size of synthesized AgNPs was 7-22 nm. The selected area diffraction (SAED) pattern and XRD spectrum revealed the crystalline structure of AgNPs. The results of FTIR analysis disclosed the functional groups responsible for the reduction of silver ion to metal nanoparticles. The biosynthesized AgNPs showed strong anti-microbial activity against drug-resistant pathogenic microorganisms. Moreover, Escherichia coli and Staphylococcus aureus were used to explore the antibacterial mechanisms of biosynthesized AgNPs. Minimal inhibitory concentrations (MICs) of E. coli and S. aureus were 6.25 µg/mL and 50 µg/mL, respectively and minimum bactericidal concentrations (MBCs) of E. coli and S. aureus were 25 µg/mL and 100 µg/mL, respectively. Results exhibited that biosynthesized AgNPs caused morphological changes and injured the membrane integrity of strains E. coli and S. aureus. The AgNPs synthesized by Sphingobium sp. MAH-11 may serve as a potent antimicrobial agent for many therapeutic applications.


Assuntos
Antibacterianos/biossíntese , Antibacterianos/farmacologia , Farmacorresistência Bacteriana/efeitos dos fármacos , Nanopartículas Metálicas/química , Prata/metabolismo , Sphingomonadaceae/metabolismo , Antibacterianos/química , Membrana Celular/efeitos dos fármacos , Escherichia coli/efeitos dos fármacos , Nanopartículas Metálicas/ultraestrutura , Testes de Sensibilidade Microbiana , Tamanho da Partícula , Filogenia , RNA Ribossômico 16S/genética , Prata/química , Prata/farmacologia , Sphingomonadaceae/classificação , Sphingomonadaceae/genética , Staphylococcus aureus/efeitos dos fármacos
7.
Environ Pollut ; 260: 114063, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32014750

RESUMO

17ß-estradiol (E2) often coexists with tetracyclines (TCs) in wastewater lagoons at intensive breeding farms, threatening the quality of surrounding water bodies. Microbial degradation is vital in E2 removal, but it is unclear how TCs affect E2 biodegradation. This primary study investigated the mechanisms of E2 degradation by Novosphingobium sp. ES2-1 in the presence of TCs and assessed the removal efficiency of E2 by strain ES2-1 in natural waters containing TCs. E2 biodegradation was unaffected at TCs concentrations below 0.1 mg L-1 yet significantly inhibited at TCs above 10 mg L-1. As elevation of TCs, E2 biodegradation rate constant decreased, and the biodegradation kinetics equation gradually deviated from the pseudo-first-order dynamics model. Importantly, the presence of TCs, especially at high-level concentrations, significantly hindered E2 ring-opening process but promoted the condensation of some phenolic ring-opening products with NH3, thereby increasing the abundance of pyridine derivatives, which were difficult to decompose over time. Additionally, strain ES2-1 could remove 52.1-100% of nature estrogens in TCs-contaminated natural waters within 7 d. Results revealed the mechanisms of TCs in E2 biodegradation and the performance of a functional strain in estrogen removal in realistic TCs-contaminated aqueous solution.


Assuntos
Estradiol/metabolismo , Sphingomonadaceae/metabolismo , Poluentes Químicos da Água/metabolismo , Biodegradação Ambiental , Estrogênios , Tetraciclinas , Águas Residuárias
8.
Ecotoxicol Environ Saf ; 192: 110254, 2020 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-32007746

RESUMO

The direct interactions of bacterial membranes and polycyclic aromatic hydrocarbons (PAHs) strongly influence the biological processes, such as metabolic activity and uptake of substrates due to changes in membrane lipids. However, the elucidation of adaptation mechanisms as well as membrane phospholipid alterations in the presence of phenanthrene (PHE) from α-proteobacteria has not been fully explored. This study was conducted to define the degradation efficiency of PHE by Sphingopyxis soli strain KIT-001 in a newly isolated from Jeonju river sediments and to characterize lipid profiles in the presence of PHE in comparison to cells grown on glucose using quantitative lipidomic analysis. This strain was able to respectively utilize 1-hydroxy-2-naphthoic acid and salicylic acid as sole carbon source and approximately 90% of PHE (50 mg/L) was rapidly degraded via naphthalene route within 1 day incubation. In the cells grown on PHE, strain KIT-001 appeared to dynamically change profiles of metabolite and lipid in comparison to cells grown on glucose. The levels of primary metabolites, phosphatidylethanolamines (PE), and phosphatidic acids (PA) were significantly decreased, whereas the levels of phosphatidylcholines (PC) and phosphatidylglycerols (PG) were significantly increased. The adaptation mechanism of Sphingopyxis sp. regarded mainly the accumulation of bilayer forming lipids and anionic lipids to adapt more quickly under restricted nutrition and toxicity condition. Hence, these findings are conceivable that strain KIT-001 has a good adaptive ability and biodegradation for PHE through the alteration of phospholipids, and will be helpful for applications for effective bioremediation of PAHs-contaminated sites.


Assuntos
Fenantrenos/metabolismo , Fosfolipídeos/metabolismo , Sphingomonadaceae/metabolismo , Biodegradação Ambiental , Sedimentos Geológicos/microbiologia , Lipidômica , Metabolômica , Naftalenos/metabolismo , Naftóis/metabolismo , Fosfolipídeos/química , Ácido Salicílico/metabolismo , Sphingomonadaceae/isolamento & purificação
9.
Molecules ; 25(1)2020 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-31906348

RESUMO

The synthesis of bioplastic from marine microbes has a great attendance in the realm of biotechnological applications for sustainable eco-management. This study aims to isolate novel strains of poly-ß-hydroxybutyrate (PHB)-producing bacteria from the mangrove rhizosphere, Red Sea, Saudi Arabia, and to characterize the extracted polymer. The efficient marine bacterial isolates were identified by the phylogenetic analysis of the 16S rRNA genes as Tamlana crocina, Bacillus aquimaris, Erythrobacter aquimaris, and Halomonas halophila. The optimization of PHB accumulation by E. aquimaris was achieved at 120 h, pH 8.0, 35 °C, and 2% NaCl, using glucose and peptone as the best carbon and nitrogen sources at a C:N ratio of 9.2:1. The characterization of the extracted biopolymer by Fourier-transform infrared spectroscopy (FTIR), Nuclear magnetic resonance (NMR), and Gas chromatography-mass spectrometry (GC-MS) proves the presence of hydroxyl, methyl, methylene, methine, and ester carbonyl groups, as well as derivative products of butanoic acid, that confirmed the structure of the polymer as PHB. This is the first report on E. aquimaris as a PHB producer, which promoted the hypothesis that marine rhizospheric bacteria were a new area of research for the production of biopolymers of commercial value.


Assuntos
Biopolímeros/biossíntese , Biopolímeros/química , Hidroxibutiratos/química , Hidroxibutiratos/metabolismo , Poliésteres/química , Poliésteres/metabolismo , Sphingomonadaceae/química , Sphingomonadaceae/metabolismo , Avicennia/microbiologia , Bacillus/química , Bacillus/genética , Bacillus/metabolismo , Biopolímeros/análise , Carbono/química , Carbono/metabolismo , Fermentação , Flavobacteriaceae/química , Flavobacteriaceae/genética , Flavobacteriaceae/metabolismo , Cromatografia Gasosa-Espectrometria de Massas , Halomonas/química , Halomonas/genética , Halomonas/metabolismo , Hidroxibutiratos/análise , Espectroscopia de Ressonância Magnética , Nitrogênio/química , Nitrogênio/metabolismo , Filogenia , Poliésteres/análise , RNA Ribossômico 16S/genética , Rizosfera , Salinidade , Arábia Saudita , Água do Mar/microbiologia , Espectroscopia de Infravermelho com Transformada de Fourier , Sphingomonadaceae/genética , Sphingomonadaceae/isolamento & purificação , Temperatura
10.
Antonie Van Leeuwenhoek ; 113(5): 719-727, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-31980980

RESUMO

In this study, a novel ginsenoside transforming bacterium, strain W1-2-3T, was isolated from mineral water. The 16S rRNA gene sequence analysis showed that strain W1-2-3T shares 93.7-92.2% sequence similarity with the members of the family Sphingomonadaceae and makes a group with Sphingoaurantiacus capsulatus YLT33T (93.7%) and S. polygranulatus MC 3718T (93.4%). The novel isolate efficiently hydrolyses the ginsenoside Rc to Rd. The genome comprises a single circular 2,880,809, bp chromosome with 3211 genes in total, and 1993 protein coding genes. The isolate was observed to grow at 10-37 °C and at pH 6-10 on R2A agar medium; maximum growth was found to occur at 25 °C and pH 7.0. Strain W1-2-3T was found to contain ubiquinone-10 as the predominant quinone and the fatty acids C16:1, C17:1ω6c, C14:0 2-OH, summed feature 3 (C16:1ω6c/C16:1ω7c) and summed feature 8 (C18:1ω6c/C18:1ω7c). The DNA G+C content was determined to be 65.9 mol%. Strain W1-2-3T can be distinguished from the other members of the family Sphingomonadaceae by a number of chemotaxonomic and phenotypic characteristics. The major polar lipids of strain W1-2-3T were identified as phosphatidylethanolamine, an unidentified glycolipid and an unidentified polar lipid. The major poly amine was found to be homospermidine. Based on polyphasic taxonomic analysis, strain W1-2-3T is concluded to represent a novel species within a new genus, for which the name Hankyongella ginsenosidimutans gen. nov., sp. nov. is proposed. The type strain of Hankyongella ginsenosidimutans is W1-2-3T (= KACC 18307T = LMG 28594T).


Assuntos
Ginsenosídeos/metabolismo , Águas Minerais/microbiologia , Sphingomonadaceae , Técnicas de Tipagem Bacteriana , DNA Bacteriano/genética , Ácidos Graxos/análise , Fosfolipídeos/análise , Filogenia , RNA Ribossômico 16S/genética , Sphingomonadaceae/classificação , Sphingomonadaceae/genética , Sphingomonadaceae/isolamento & purificação , Sphingomonadaceae/metabolismo , Ubiquinona/análogos & derivados , Ubiquinona/análise
11.
Colloids Surf B Biointerfaces ; 185: 110633, 2020 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-31740324

RESUMO

The microcystin-LR (MC-LR) produced due to harmful cyanobacterial blooms have brought great harm to human and aquatic organisms, attracting a wide public health attention. To deal with MC-LR contamination, we synthesized a novel bio-functionalized composite for the high-efficient and sustainable biodegradation of microcystin-LR by covalent immobilizing Sphingopyxis sp. YF1 onto chitosan-grafted Fe3O4 magnetic particles (Fe3O4@CTS). The Fourier transform infrared spectroscopy (FTIR), Field emission scanning electron microscopy (SEM) and vibrating sample magnetometry (VSM) were utilized to characterize the structural properties of Fe3O4@CTS/Sphingopyxis sp. YF1. The immobilization conditions were optimized. And the MC-LR-degrading capabilities of Fe3O4@CTS/Sphingopyxis sp. YF1 were assessed under various conditions. The results showed that the optimal immobilization conditions containing 1.0 % (v/v) glutaraldehyde, immobilization for 4 h at 30 ℃. The Fe3O4@CTS/Sphingopyxis sp. YF1 showed an attractive degradation performance which possesed a wide torlerance to pH (6.0-9.0) and temperature (25-35 ℃). More interesting is that the Fe3O4@CTS/Sphingopyxis sp. YF1 exhibited significantly increased MC-LR-degrading capabilities after recycling and reusing which degradation rate reached 1.50 µg/mL/h in the sixth cycle, and it was easily recycled by using a magnet (Ms 21.5 emug-1). Two intermediates (tetrapeptide and Adda) and three degradation related genes (mlrA, mlrB and mlrC) were obtained in this study and the pathway for the degradation was proposed. These results revealed that Fe3O4@CTS/Sphingopyxis sp. YF1 can be potentially used for treatment of MC-LR contaminated environment.


Assuntos
Quitosana/química , Compostos Férricos/química , Microcistinas/metabolismo , Sphingomonadaceae/metabolismo , Biodegradação Ambiental , Células Imobilizadas/citologia , Glutaral/química , Concentração de Íons de Hidrogênio , Toxinas Marinhas , Sphingomonadaceae/citologia , Sphingomonadaceae/ultraestrutura , Temperatura
12.
Ecotoxicol Environ Saf ; 187: 109848, 2020 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-31670182

RESUMO

Dimethyl terephthalate (DMT) is a primary ingredient widely used in the manufacture of polyesters and industrial plastics; its environmental fate is of concern due to its global use. Microorganisms play key roles in the dissipation of DMT from the environment; however, the enzymes responsible for the initial transformation of DMT and the possible altered toxicity due to this biotransformation have not been extensively studied. To reduce DMT toxicity, we identified the esterase gene dmtH involved in the initial transformation of DMT from the AOPP herbicide-transforming strain Sphingobium sp. C3. DmtH shows 24-41% identity with α/ß-hydrolases and belongs to subfamily V of bacterial esterases. The purified recombinant DmtH was capable of transforming DMT to mono-methyl terephthalate (MMT) and potentially transforming other p-phthalic acid esters, including diallyl terephthalate (DAT) and diethyl terephthalate (DET). Using C. elegans as an assay model, we observed the severe toxicity of DMT in inducing reactive oxygen species (ROS) production, decreasing locomotion behavior, reducing lifespan, altering molecular basis for oxidative stress, and inducing mitochondrial stress. In contrast, exposure to MMT did not cause obvious toxicity, induce oxidative stress, and activate mitochondrial stress in nematodes. Our study highlights the usefulness of Sphingobium sp. C3 and its esterase DmtH in transforming p-phthalic acid esters and reducing the toxicity of DMT to organisms.


Assuntos
Poluentes Ambientais/toxicidade , Esterases/genética , Genes Bacterianos , Ácidos Ftálicos/toxicidade , Sphingomonadaceae/metabolismo , Animais , Biodegradação Ambiental , Biotransformação , Caenorhabditis elegans/efeitos dos fármacos , Caenorhabditis elegans/metabolismo , Poluentes Ambientais/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Ácidos Ftálicos/metabolismo , Plásticos/química , Sphingomonadaceae/enzimologia , Sphingomonadaceae/genética
13.
J Toxicol Environ Health A ; 82(22): 1164-1171, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31833448

RESUMO

The aromatic compound phenylacetic acid (PAA) is present in the environment, and released in the catabolism of phenylalanine, 2-phenylethylamine, or environmental contaminants such as ethylbenzene and styrene. PAA was also proposed to be involved in human chronic kidney disease development. Several bacteria and fungi utilize these aromatic acids as sole carbon source either during aerobic or anaerobic conditions. The aromatic structure of PAA makes this compound resistant toward oxidation or reduction, because the stabilizing resonance energy of the aromatic ring system is difficult to overcome. In the case of bacteria that utilize aromatic compounds as growth substrates, the aromatic ring system limits survival due to a lack of carbon source. Sphingopyxis sp. YF1 isolated from Lake Taihu was found to be beneficial in bioremediation of aromatic compounds. This study thus aimed to examine the influence of environmental factors such as temperature, PAA concentration, and pH on the effectiveness of Sphingopyxis sp. YF1 to degrade aromatic compounds using PAA as model compound. Data showed the highest PAA-degrading rate of strain Sphingopyxis sp. YF1 was 7.6 mg/L·h under the condition of 20°C, pH 9 with a 1000 µg/ml concentration of PAA. Evidence indicates that PAA-degrading ability of strain Sphingopyxis sp. YF1 appears to be primarily influenced by the concentration of PAA, followed by temperature and pH. PAA-degrading gene PAAase was identified in this strain using polymerase chain reaction (PCR) method. These results illustrate that the bacteria Sphingopyxis sp. YF1 removes PAA effectively at certain environmental conditions and this proves beneficial in bioremediation of aromatic compounds.


Assuntos
Biodegradação Ambiental , Hidrocarbonetos Aromáticos/metabolismo , Lagos/química , Fenilacetatos/metabolismo , Sphingomonadaceae/metabolismo , Poluentes Químicos da Água/metabolismo , China
14.
J Agric Food Chem ; 67(44): 12228-12236, 2019 Nov 06.
Artigo em Inglês | MEDLINE | ID: mdl-31638826

RESUMO

Zeaxanthin is a value-added carotenoid with wide applications. This study aims to manipulate a generally recognized as safe and carotenoid-producing bacterium, Sphingobium sp., for enhanced production of zeaxanthin and exopolysaccharides. First, whole-genome sequencing and analysis of pathway genes were applied to define the carotenoid pathway in Sphingobium sp. Second, a Sphingobium transformation system was established to engineer metabolite flux into zeaxanthin. By a combination of chemical mutagenesis and removal of bottlenecks of carotenoid biosynthesis via overexpression of three rate-limiting enzymes, the genetically modified Sphingobium DIZ strain produced 21.26 mg/g dry cell weight of zeaxanthin, which was about 4-fold higher than the wild type. Upon optimization of culture conditions, the DIZ strain produced 479.5 mg/L of zeaxanthin with the productivity of 4.99 mg/L/h and 21.9 g/L of exopolysaccharides using a fed-batch fermentation strategy. This study represents the first genetic manipulation of Sphingobium sp., a biotechnologically important bacterium, for high-yield production of value-added metabolites.


Assuntos
Proteoglicanas/biossíntese , Sphingomonadaceae/genética , Sphingomonadaceae/metabolismo , Zeaxantinas/biossíntese , Técnicas de Cultura Celular por Lotes , Meios de Cultura/metabolismo , Fermentação , Engenharia Metabólica
15.
Genes (Basel) ; 10(9)2019 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-31500174

RESUMO

Sphingopyxis inhabit diverse environmental niches, including marine, freshwater, oceans, soil and anthropogenic sites. The genus includes 20 phylogenetically distinct, valid species, but only a few with a sequenced genome. In this work, we analyzed the nearly complete genome of the newly described species, Sphingopyxis lindanitolerans, and compared it to the other available Sphingopyxis genomes. The genome included 4.3 Mbp in total and consists of a circular chromosome, and two putative plasmids. Among the identified set of lin genes responsible for γ-hexachlorocyclohexane pesticide degradation, we discovered a gene coding for a new isoform of the LinA protein. The significant potential of this species in the remediation of contaminated soil is also correlated with the fact that its genome encodes a higher number of enzymes potentially involved in aromatic compound degradation than for most other Sphingopyxis strains. Additional analysis of 44 Sphingopyxis representatives provides insights into the pangenome of Sphingopyxis and revealed a core of 734 protein clusters and between four and 1667 unique proteins per genome.


Assuntos
Genoma Bacteriano , Hexaclorocicloexano/metabolismo , Praguicidas/metabolismo , Sphingomonadaceae/genética , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Biodegradação Ambiental , Sphingomonadaceae/enzimologia , Sphingomonadaceae/metabolismo
16.
Toxins (Basel) ; 11(10)2019 09 20.
Artigo em Inglês | MEDLINE | ID: mdl-31547007

RESUMO

Biodegradation is efficient for removing cyanobacterial toxins, such as microcystins (MCs) and nodularin (NOD). However, not all the microbial strains with the microcystin-biodegrading enzymes MlrA and MlrC could biodegrade NOD. Studies on genes and enzymes for biodegrading NOD can reveal the function and the biodegradation pathway of NOD. Based on successful cloning and expression of the USTB-05-A and USTB-05-C genes from Sphingopyxis sp. USTB-05, which are responsible for the biodegradation of MCs, the pathway for biodegrading NOD by these two enzymes was investigated in this study. The findings showed that the enzyme USTB-05-A converted cyclic NOD (m/z 825.4516) into its linear type as the first product by hydrolyzing the arginine and Adda peptide bond, and that USTB-05-C cut off the Adda and glutamic acid peptide bond of linearized NOD (m/z 843.4616) and produced dimeric Adda (m/z 663.4377) as the second product. Further, based on the homology modeling of enzyme USTB-05-A, site-directed mutants of USTB-05-A were constructed and seven crucial sites for enzyme USTB-05-A activity were found. A complete enzymatic mechanism for NOD biodegradation by USTB-05-A in the first step was proposed: glutamic acid 172 and histidine 205 activate a water molecule facilitating a nucleophilic attack on the arginine and Adda peptide bond of NOD; tryptophan 176 and tryptophan 201 contact the carboxylate side chain of glutamic acid 172 and accelerate the reaction rates; and histidine 260 and asparagine 264 function as an oxyanion hole to stabilize the transition states.


Assuntos
Peptídeos Cíclicos/metabolismo , Sphingomonadaceae/metabolismo , Biodegradação Ambiental , Clonagem Molecular , Redes e Vias Metabólicas , Peptídeos Cíclicos/química , Sphingomonadaceae/genética
17.
J Hazard Mater ; 380: 120881, 2019 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-31307001

RESUMO

Three bacterial consortia, named YC-SY1, YC-BJ1 and YC-GZ1, were enriched from different areas of China. Bacterial consortia YC-SY1, YC-BJ1 and YC-GZ1 could efficiently degrade triphenyl phosphate (TPhP) (100 mg/L) by approximately 79.4%, 99.8% and 99.6%, tricresyl phosphate (TCrP) by 90.6%, 91.9% and 96.3%, respectively, within 4 days. And they could retain high degrading efficiency under a broad range of temperature (15-40 ℃), pH (6.0-10.0) and salinity (0-4%). A total of 10 bacterial isolates were selected and investigated their degradation capacity. Among these isolates, two were significantly superior to the others. Strain Rhodococcus sp. YC-JH2 could utilize TPhP (50 mg/L) as sole carbon source for growth with 37.36% degradation within 7 days. Strain Sphingopyxis sp. YC-JH3 could efficiently degrade 96.2% of TPhP (50 mg/L) within 7 days, except that no cell growth was observed. Combined with 16S diversity analysis, our results suggest that the effective components of three bacterial consortia responsible for TPhP and TCrP degradation were almost the same, that is, bacteria capable of degrading TPhP and TCrP are limited, in this study, the most efficient component is Sphingopyxis. This study provides abundant microorganism sources for research on organophosphorus flame retardants (OPFRs) metabolism and bioremediation towards OPFRs-contaminated environments.


Assuntos
Retardadores de Chama/metabolismo , Metagenômica , Consórcios Microbianos , Compostos Organofosforados/metabolismo , Rhodococcus/metabolismo , Sphingomonadaceae/metabolismo
18.
Environ Sci Pollut Res Int ; 26(25): 25932-25944, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31273663

RESUMO

A natural phenanthrene-degrading consortium CON was inoculated with an exogenous strain Sphingobium sp. (ex Sp. paucimobilis) 20006FA yielding the consortium called I-CON, in order to study ecological interactions into the bacterial community. DGGE and proteomic profiles and analyses by HTS (High-Throughput Sequencing) technologies demonstrated inoculant establishment and changes on CON composition. Inoculation increased degradation efficiency in I-CON and prevented intermediate HNA accumulation. This could be explained not only by the inoculation, but also by enrichment in Achromobacter genus at expense of a decrease in Klebsiella genus. After inoculation, cooperation between Sphingobium and Achromobacter genera were improved, thereby, some competition could have been generated, and as a consequence, species in minor proportion (cheaters), as Inquilinus sp. and Luteibacter sp., were not detected. Sequences of Sphingobium (corresponding to the inoculated strain) did not vary. PICRUSt predicted a network with bacterial phylotypes connected with enzymes, showing functional redundancy in the phenanthrene pathway, with exception of the first enzymes biphenyl-2,3-diol 1,2-dioxygenase and protocatechuate 4,5-dioxygenase that were only encoded in Sphingobium sp. This is the first report where a natural consortium that has been characterized by HTS technologies is inoculated with an exogenous strain in order to study competitiveness and interactions.


Assuntos
Achromobacter/química , Achromobacter/metabolismo , Dioxigenases/metabolismo , Fenantrenos/química , Proteômica/métodos , Sphingomonadaceae/metabolismo , Biodegradação Ambiental , Dioxigenases/química , Sphingomonadaceae/química
19.
J Bacteriol ; 201(17)2019 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-31209079

RESUMO

The herbicide dicamba is initially degraded via the tetrahydrofolate (THF)-dependent demethylation system in Rhizorhabdus dicambivorans Ndbn-20. Two THF-dependent dicamba methyltransferase gene clusters, scaffold 50 and scaffold 66, were found in the genome of strain Ndbn-20. Each cluster contains a dicamba methyltransferase gene and three THF metabolism-related genes, namely, metF (coding for 5,10-CH2-THF reductase), folD (coding for 5,10-CH2-THF dehydrogenase-5,10-methenyl-THF cyclohydrolase), and purU (coding for 10-formyl-THF deformylase). In this study, reverse transcription-PCR (RT-PCR) results showed that only genes in scaffold 66, not those in scaffold 50, were transcribed in dicamba-cultured cells. The metF gene of scaffold 66 (metF1) was expressed in Escherichia coli BL21(DE3), and the product was purified as a His6-tagged protein. Purified MetF1 was found to be a monomer and exhibited 5-CH3-THF dehydrogenase activity in vitro The k cat and Km for 5-CH3-THF were 0.23 s-1 and 16.48 µM, respectively. However, 5,10-CH2-THF reductase activity was not detected for MetF1 under the conditions tested. Gene disruption results showed that metF1 is essential for dicamba degradation, whereas folD1 is dispensable.IMPORTANCE There are several THF-dependent methyltransferase genes and THF-metabolic genes in the genome of R. dicambivorans Ndbn-20; however, which genes are involved in dicamba demethylation and the mechanism underlying THF regeneration remain unknown. This study revealed that scaffold 66 is responsible for dicamba demethylation and that MetF1 physiologically catalyzes the dehydrogenation of 5-CH3-THF to 5,10-CH2-THF in the THF-dependent dicamba demethylation system in R. dicambivorans Ndbn-20. Furthermore, the results showed that MetF1 differs from previously characterized MetF in phylogenesis, biochemical properties, and catalytic activity; e.g., MetF1 in vitro did not show 5,10-CH2-THF reductase activity, which is the physiological function of Escherichia coli MetF. This study provides new insights into the mechanism of the THF-dependent methyltransferase system.


Assuntos
Proteínas de Bactérias/metabolismo , Dicamba/metabolismo , Oxirredutases/metabolismo , Sphingomonadaceae/enzimologia , Tetra-Hidrofolatos/metabolismo , Proteínas de Bactérias/genética , Desmetilação , Regulação Bacteriana da Expressão Gênica , Regulação Enzimológica da Expressão Gênica , Oxirredutases/genética , Filogenia , Sphingomonadaceae/genética , Sphingomonadaceae/metabolismo
20.
Chemosphere ; 234: 789-795, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31247488

RESUMO

Grasses are advantageous in the removal of polycyclic aromatic hydrocarbons (PAHs) in soil because of their fibrous root, high tolerance to environmental stress, and low nutritional requirements. In this study, a pot experiment was conducted to test the ability of four grasses to remove PAHs in the soil, and to investigate the corresponding bacterial community shift in the rhizosphere of each. Sudangrass achieved the maximum removal of PAHs at 98% dissipation rate after 20 days. Polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) and next-generation sequencing revealed that sudangrass specially enriched the growth of a known PAHs degrader, Sphingomonadales, regardless of the presence or absence of PAHs in the soil. Moreover, the gene copy numbers of PAHs catabolic genes, PAH-RHDα and nidA, as measured by real time-PCR (RT-PCR) were highest in the soil planted with sudangrass. Overall, this study suggested that sudangrass further enhanced the dissipation of PAHs by enriching Sphingomonadales in its rhizosphere.


Assuntos
Biodegradação Ambiental , Hidrocarbonetos Policíclicos Aromáticos/metabolismo , Rizosfera , Sorghum/microbiologia , Bactérias/isolamento & purificação , Bactérias/metabolismo , Poaceae/metabolismo , Hidrocarbonetos Policíclicos Aromáticos/análise , Poluentes do Solo/análise , Poluentes do Solo/metabolismo , Sphingomonadaceae/crescimento & desenvolvimento , Sphingomonadaceae/metabolismo
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