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1.
Ecotoxicol Environ Saf ; 195: 110375, 2020 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-32200142

RESUMO

Microbial immobilization is a novel and environmentally friendly technology that uses microbes to reduce metal availability in soil and accumulation of heavy metals in plants. We used urea agar plates to isolate urease-producing bacteria from the rhizosphere soil of pakchoi in Cd- and Pb-contaminated farmland and investigated their effects on Cd and Pb accumulation in pakchoi and the underlying mechanisms. The results showed that two urease-producing bacteria, Bacillus megaterium N3 and Serratia liquefaciens H12, were identified by screening. They had higher ability to produce urease (57.5 ms cm-1 min-1 OD600-1 and 76.4 ms cm-1 min-1 OD600-1, respectively). The two strains allowed for the immobilization of Cd and Pb by extracellular adsorption, bioprecipitation, and increasing the pH (from 6.94 to 7.05-7.09), NH4+ content (69.1%-127%), and NH4+/NO3- ratio (from 1.37 to 1.67-2.11), thereby reducing the DTPA-extractable Cd (35.3%-58.8%) and Pb (37.8%-62.2%) contents in the pakchoi rhizosphere soils and the Cd (76.5%-79.7%) and Pb (76.3%-83.5%) contents in the leaves (edible tissue) of pakchoi. The strains were highly resistant to heavy metal toxicity; produced IAA, siderophores and abscisic acid; and increased the NH4+/NO3- ratio, which might be related to the two strains protectiing pakchoi against the toxic effect of Cd and Pb and increasing pakchoi biomass. Thus, the results were supposed to strain resources and a theoretical basis for the remediation of Cd- and Pb-contaminated farmlands for the safe production of vegetables.


Assuntos
Bacillus megaterium/isolamento & purificação , Brassica/crescimento & desenvolvimento , Cádmio/análise , Chumbo/análise , Serratia liquefaciens/isolamento & purificação , Microbiologia do Solo , Poluentes do Solo/análise , Bacillus megaterium/metabolismo , Biodegradação Ambiental , Biomassa , Brassica/metabolismo , Cádmio/metabolismo , Fazendas , Chumbo/metabolismo , Rizosfera , Serratia liquefaciens/metabolismo , Solo/química , Poluentes do Solo/metabolismo , Urease/metabolismo
2.
Arch Microbiol ; 202(6): 1381-1396, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32179939

RESUMO

Plant growth-promoting rhizobacteria (PGPR) are widely used to improve plant nutrient uptake and assimilation and soil physicochemical properties. We investigated the effects of bacterial (Bacillus megaterium strain DU07) fertilizer applications in a eucalyptus (clone DH32-29) plantation in Guangxi, China in February 2011. We used two types of organic matter, i.e., fermented tapioca residue ("FTR") and filtered sludge from a sugar factory ("FS"). The following treatments were evaluated: (1) no PGPR and no organic matter applied (control), (2) 3 × 109 CFU/g (colony forming unit per gram) PGPR plus FS (bacterial fertilizer 1, hereafter referred to as BF1), (3) 4 × 109 CFU/g plus FS (BF2), (4) 9 × 109 CFU/g plus FS (BF3), (5) 9 × 109 CFU/g broth plus FTR (BF4). Soil and plant samples were collected 3 months (M3) and 6 months (M6) after the seedlings were planted. In general, bacterial fertilizer amendments significantly increased plant foliar total nitrogen (TN) and soil catalase activity in the short term (month 3, M3); whereas, it significantly increased foliar TN, chlorophyll concentration (Chl-ab), proline; plant height, diameter, and volume of timber; and soil urease activity, STN, and available N (Avail N) concentrations in the long term (month 6, M6). Redundancy analysis showed that soil available phosphorus was significantly positively correlated with plant growth in M3, and soil Avail N was negatively correlated with plant growth in M6. In M3, soil catalase was more closely correlated with plant parameters than other enzyme activities and soil nutrients, and in M6, soil urease, polyphenol oxidase, and peroxidase were more closely correlated with plant parameters than other environmental factors and soil enzyme activities. PCA results showed that soil enzyme activities were significantly improved under all treatments relative to the control. Hence, photosynthesis, plant growth, and soil N retention were positively affected by bacterial fertilizer in M6, and bacterial fertilizer applications had positive and significant influence on soil enzyme activities during the trial period. Thus, bacterial fertilizer is attractive for use as an environmentally friendly fertilizer in Eucalyptus plantations following proper field evaluation.


Assuntos
Bacillus megaterium/metabolismo , Eucalyptus/crescimento & desenvolvimento , Fertilizantes/microbiologia , Plântula/crescimento & desenvolvimento , Solo/química , Catalase/metabolismo , China , Clorofila/análise , Fertilizantes/análise , Manihot/microbiologia , Nitrogênio/análise , Nutrientes , Fósforo/análise , Desenvolvimento Vegetal , Esgotos/microbiologia , Microbiologia do Solo , Urease/metabolismo
3.
J Basic Microbiol ; 60(1): 22-26, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31692013

RESUMO

Metals are among the most prevalent pollutants released into the environment. For these reasons, the use of biomarkers for environmental monitoring of individuals and populations exposed to metal pollution has gained considerable attention, offering fast and sensitive detection of chemical stress in organisms. There are different metal resistance genes in bacteria that can be used as biomarkers, including cation diffusion facilitators carrying metal ions; the prototype is the cobalt-zinc-cadmium transporter (czcD). The present study reports the expression changes in the czcD gene in Bacillus megaterium and Microbacterium liquefaciens under nickel and vanadium exposure by real-time polymerase chain reaction. The nickel-vanadium-resistant strains of B. megaterium and M. liquefaciens used in this study were isolated from mine tailings in Guanajuato, Mexico. The czcD gene showed high expression under exposure to 200 ppm of Ni and 200 ppm of V during the logarithmic growth phase of M. liquefaciens in PHGII liquid media. In contrast, no changes were observed in B. megaterium during logarithmic and stationary growth, perhaps due to the gene having differential expression during the growth phases. The expression profiles obtained for czcD show the possibility of using this gene from M. liquefaciens as a biomarker of nickel and vanadium pollution in microorganisms.


Assuntos
Actinobacteria/genética , Bacillus megaterium/genética , Biomarcadores Ambientais/genética , Genes Bacterianos/genética , Actinobacteria/metabolismo , Bacillus megaterium/metabolismo , Expressão Gênica , México , Mineração , Níquel/metabolismo , Microbiologia do Solo , Poluentes do Solo/metabolismo , Vanádio/metabolismo
4.
An Acad Bras Cienc ; 91(3): e20180694, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31618412

RESUMO

The strains CM-Z19 and CM-Z6, which are capable of highly degrading chlorpyrifos-methyl, were isolated from soil. They were identified as Bacillus megaterium CM-Z19 and Pseudomonas syringae CM-Z6, respectively, based on the 16S rRNA and an analysis of their morphological, physiological and biochemical characteristics. The strain CM-Z19 showed 92.6% degradation of chlorpyrifos-methyl (100 mg/L) within 5 days of incubation, and the strain CM-Z6 was 99.1% under the same conditions. In addition, the degradation characteristics of the two strains were compared and studied, and the results showed that the strain CM-Z19 had higher phosphoesterase activity and ability to degrade the organophosphorus pesticide than did the strain CM-Z6. However, the strain CM-Z19 could not degrade its first hydrolysis metabolite 3,5,6-trichloro-2-pyridinol (TCP) and could not completely degrade chlorpyrifos-methyl. The strain CM-Z6 could effectively degrade TCP and could degrade chlorpyrifos-methyl more quickly than strain CM-Z19.


Assuntos
Bacillus megaterium/metabolismo , Biodegradação Ambiental , Clorpirifos/análogos & derivados , Praguicidas/metabolismo , Pseudomonas syringae/metabolismo , Bacillus megaterium/isolamento & purificação , Clorpirifos/isolamento & purificação , Clorpirifos/metabolismo , Inseticidas/isolamento & purificação , Inseticidas/metabolismo , Praguicidas/isolamento & purificação , Pseudomonas syringae/isolamento & purificação , RNA Ribossômico 16S/metabolismo , Microbiologia do Solo
5.
J Chem Ecol ; 45(9): 789-797, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31422514

RESUMO

Peritassa campestris (Celastraceae) root bark accumulates potent antitumor quinonemethide triterpenes (QMTs). When grown in their natural habitat, plants of the family Celastraceae produce different QMTs such as celastrol (3) and pristimerin (4). However, when they are inserted in in vitro culture systems, they accumulate maytenin (1) as the main compound. Recently, Bacillus megaterium was detected as an endophytic microorganism (EM) living inside P. campestris roots cultured in vitro. We hypothesized that compound (1) controls EM growth more efficiently, and that the presence of EMs in the root culture causes compound (1) to accumulate. For the first time, this work has explored plant-microorganism interaction in a species of the family Celastraceae by co-culture with an EM. Live endophytic bacteria were used, and QMT accumulation in P. campestris adventitious roots was our main focus. The antimicrobial activity of the main QMTs against endophytic B. megaterium was also evaluated. Our results showed that compound (1) and maytenol (5) were more effective than their precursors QMTs (3) and (4) in controlling the EM. Co-culture of B. megaterium with roots significantly reduced bacterial growth whereas root development remained unaffected. Compound (1) production was 24 times higher after 48 hr in the presence of the highest B. megaterium concentration as compared to the control. Therefore, P. campestris adventitious roots affect the development of the endophyte B. megaterium through production of QMTs, which in turn can modulate production of compound (1).


Assuntos
Bacillus megaterium/metabolismo , Celastraceae/metabolismo , Extratos Vegetais/química , Raízes de Plantas/metabolismo , Triterpenos/química , Anti-Infecciosos/química , Antineoplásicos/química , Endófitos/metabolismo , Fatores de Tempo
6.
Microb Cell Fact ; 18(1): 132, 2019 Aug 12.
Artigo em Inglês | MEDLINE | ID: mdl-31405374

RESUMO

BACKGROUND: Heparosan is the unsulfated precursor of heparin and heparan sulfate and its synthesis is typically the first step in the production of bioengineered heparin. In addition to its utility as the starting material for this important anticoagulant and anti-inflammatory drug, heparosan is a versatile compound that possesses suitable chemical and physical properties for making a variety of high-quality tissue engineering biomaterials, gels and scaffolds, as well as serving as a drug delivery vehicle. The selected production host was the Gram-positive bacterium Bacillus megaterium, which represents an increasingly used choice for high-yield production of intra- and extracellular biomolecules for scientific and industrial applications. RESULTS: We have engineered the metabolism of B. megaterium to produce heparosan, using a T7 RNA polymerase (T7 RNAP) expression system. This system, which allows tightly regulated and efficient induction of genes of interest, has been co-opted for control of Pasteurella multocida heparosan synthase (PmHS2). Specifically, we show that B. megaterium MS941 cells co-transformed with pT7-RNAP and pPT7_PmHS2 plasmids are capable of producing heparosan upon induction with xylose, providing an alternate, safe source of heparosan. Productivities of ~ 250 mg/L of heparosan in shake flasks and ~ 2.74 g/L in fed-batch cultivation were reached. The polydisperse Pasteurella heparosan synthase products from B. megaterium primarily consisted of a relatively high molecular weight (MW) heparosan (~ 200-300 kD) that may be appropriate for producing certain biomaterials; while the less abundant lower MW heparosan fractions (~ 10-40 kD) can be a suitable starting material for heparin synthesis. CONCLUSION: We have successfully engineered an asporogenic and non-pathogenic B. megaterium host strain to produce heparosan for various applications, through a combination of genetic manipulation and growth optimization strategies. The heparosan products from B. megaterium display a different range of MW products than traditional E. coli K5 products, diversifying its potential applications and facilitating increased product utility.


Assuntos
Bacillus megaterium/genética , Bacillus megaterium/metabolismo , Dissacarídeos/biossíntese , Glicosiltransferases/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Vias Biossintéticas , RNA Polimerases Dirigidas por DNA/genética , Engenharia Genética , Glicosiltransferases/genética , Engenharia Metabólica , Pasteurella multocida/enzimologia , Proteínas Virais/genética
7.
Biotechnol Lett ; 41(8-9): 951-961, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31278569

RESUMO

OBJECTIVES: A three-species consortium for one-step fermentation of 2-keto-L-gulonic acid (2-KGA) was constructed to better strengthen the cell-cell communication. And the programmed cell death module based on the LuxI/LuxR quorum-sensing (QS) system was established in Gluconobacter oxydans to reduce the competition that between G. oxydans and Ketogulonicigenium vulgare. RESULTS: By constructing and optimizing the core region of the promoter, which directly regulated the expression of lethal ccdB genes in QS system, IR3C achieved the best lethal effect. The consortium of IR3C- K. vulgare-Bacillus megaterium (abbreviated as 3C) achieved the highest 2-KGA titer (68.80 ± 4.18 g/l), and the molar conversion rate was 80.7% within 36 h in 5 l fermenter. Metabolomic analysis on intracellular small molecules of consortia 3C and 1C showed that most amino acids (such as glycine, leucine, methionine and proline) and TCA cycle intermediates (such as succinic acid, fumaric acid and malic acid) were significantly affected. These results further validated that the programmed cell death module based on the LuxI/LuxR QS system in G. oxydans could also faciliate better growth and higher production of consortium 3C for one-step fermentation. CONCLUSIONS: We successfully constructed a novel three-species consortia for one-step vitamin C fermentation by strengthening the cell-cell communication. This will be very useful for probing the rational design principles of more complex multi-microbial consortia.


Assuntos
Ácido Ascórbico/metabolismo , Bacillus megaterium/metabolismo , Fermentação , Gluconobacter oxydans/metabolismo , Consórcios Microbianos , Rhodobacteraceae/metabolismo , Açúcares Ácidos/metabolismo , Bacillus megaterium/crescimento & desenvolvimento , Comunicação Celular , Gluconobacter oxydans/crescimento & desenvolvimento , Interações Microbianas , Rhodobacteraceae/crescimento & desenvolvimento , Vitaminas/metabolismo
8.
Artigo em Inglês | MEDLINE | ID: mdl-31324006

RESUMO

The inoculation of beneficial microorganisms to improve plant growth and soil properties is a promising strategy in the soil amendment. However, the effects of co-inoculation with phosphate-solubilizing bacteria (PSB) and N2-fixing bacteria (NFB) on the soil properties of typical C-deficient soil remain unclear. Based on a controlled experiment and a pot experiment, we examined the effects of PSB (M: Bacillus megaterium and F: Pseudomonas fluorescens), NFB (C: Azotobacter chroococcum and B: Azospirillum brasilence), and combined PSB and NFB treatments on C, N, P availability, and enzyme activities in sterilized soil, as well as the growth of Cyclocarya Paliurus seedlings grow in unsterilized soil. During a 60-day culture, prominent increases in soil inorganic N and available P contents were detected after bacteria additions. Three patterns were observed for different additions according to the dynamic bacterial growth. Synergistic effects between NFB and PSB were obvious, co-inoculations with NFB enhanced the accumulation of available P. However, decreases in soil available P and N were observed on the 60th day, which was induced by the decreases in bacterial quantities under C deficiency. Besides, co-inoculations with PSB and NFB resulted in greater performance in plant growth promotion. Aimed at amending soil with a C supply shortage, combined PSB and NFB treatments are more appropriate for practical fertilization at intervals of 30-45 days. The results demonstrate that co-inoculations could have synergistic interactions during culture and application, which may help with understanding the possible mechanism of soil amendment driven by microorganisms under C deficiency, thereby providing an alternative option for amending such soil.


Assuntos
Fixação de Nitrogênio , Nitrogênio/metabolismo , Fósforo/metabolismo , Microbiologia do Solo , Solo/química , Azospirillum brasilense/metabolismo , Azotobacter/metabolismo , Bacillus megaterium/metabolismo , Carga Bacteriana , China , Pseudomonas fluorescens/metabolismo
9.
Curr Microbiol ; 76(10): 1215-1224, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31254008

RESUMO

Nejayote is an alkaline wastewater generated during the nixtamalization process. Nejayote contains high-value compounds such as ferulic acid (FA), which is widely employed as a substrate for the biotechnological production of flavors and aromas. In the present study, the isolation, identification, and characterization of a native strain of Bacillus megaterium were performed, and its capacity to produce 4-vinylguaiacol (4VG) from ferulic acid was evaluated by employing growing cell and resting cell systems. Growing cells of native B. megaterium biotransformed 6 mM crude FA in nejayote into 2.1 mM 4VG, reaching a productivity of 0.21 mM h-1 4VG, while nejayote enriched with FA at 10, 15, and 25 mM resulted in the formation of 2.4, 3.8, and 6.2 mM 4VG and productivities of 0.24, 0.38, and 0.51 mM h-1 4VG, respectively. In the resting cell system, from 6 and 25 mM pure FA, 3.5 mM 4VG was produced (0.18 mM h-1 4VG), while at 10 and 15 mM FA, 4.6 and 5.1 mM 4VG (average of 0.24 mM h-1 4VG) were obtained, respectively. The native B. megaterium strain, isolated from nejayote, showed great biotechnological potential to produce 4VG from crude FA contained in this wastewater, in which other Bacillus species, such as B. licheniformis and B. cereus, were unable to grow and biotransform FA into 4VG.


Assuntos
Bacillus megaterium/classificação , Bacillus megaterium/metabolismo , Ácidos Cumáricos/metabolismo , Águas Residuárias/microbiologia , Zea mays , Bacillus megaterium/genética , Bacillus megaterium/crescimento & desenvolvimento , Biomassa , Biotransformação , Ácidos Cumáricos/química , Guaiacol/análogos & derivados , Guaiacol/metabolismo , Filogenia , Águas Residuárias/química
10.
Appl Microbiol Biotechnol ; 103(17): 7191-7202, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31250062

RESUMO

A low production rate for calcium carbonate with microbial solidification technology at low temperatures often restricts its application. For this reason, adding urea to the medium and the domestication of Bacillus megaterium at low temperature were proposed to produce more calcium carbonate based on an analysis of growth characteristics, urease activity, and the production rates for calcium carbonate under different conditions. Sand solidification tests were conducted to demonstrate improvements caused by the methods. The results showed that the higher the temperature, the faster the growth of Bacillus megaterium and the stronger the urease activity. Growth was fastest and urease activity strongest at a pH of 8. Adding urea to the medium and the domestication of B. megaterium at low temperature can both improve the production rate, effectively increasing calcium carbonate precipitation at low temperature. Combining the two methods resulted in greater improvement of the production rate for calcium carbonate. The two methods were also found to improve the effect of sand solidification. Therefore, our study provides a solid foundation for the actual engineering application of bio-cementation technology at low temperature.


Assuntos
Bacillus megaterium/crescimento & desenvolvimento , Bacillus megaterium/metabolismo , Biomineralização , Carbonato de Cálcio/metabolismo , Bacillus megaterium/enzimologia , Carbonato de Cálcio/química , Precipitação Química , Temperatura Baixa , Materiais de Construção/microbiologia , Meios de Cultura/química , Concentração de Íons de Hidrogênio , Ureia/análise , Ureia/metabolismo , Urease/metabolismo
11.
Carbohydr Polym ; 219: 130-142, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31151510

RESUMO

The physicochemical properties and biological activity of levan, a generic term given to oligo- and polysaccharides consisting of fructose units linked predominantly by ß(2-6) bonds, are attributable to both its size and structural complexity. Branching in ß(2-1) contributes to diversify levan structures and properties. There is a broad spectrum of applications for levan and accordingly it has been the subject of several comprehensive reviews. A thorough analysis focused on the product specificity of enzymes from the Glycoside-Hydrolase family 68 that synthesize levan is however missing. We analyze here traditional and novel strategies to manipulate bacterial levansucrases in favor of the generation of low- or high-molecular weight levan, including site directed mutagenesis and chemical engineering. A comparison of highly variable structural elements of levansucrases is presented in the context of their capacity to synthesize saccharides of different sizes, employing the levansucrases from Bacillus subtilis and Bacillus megaterium as references.


Assuntos
Bacillus megaterium/metabolismo , Bacillus subtilis/metabolismo , Frutanos , Hexosiltransferases/genética , Engenharia Química , Frutanos/química , Frutanos/metabolismo , Peso Molecular , Mutagênese Sítio-Dirigida
12.
Proc Natl Acad Sci U S A ; 116(23): 11470-11479, 2019 06 04.
Artigo em Inglês | MEDLINE | ID: mdl-31113879

RESUMO

Germination of Bacillus spores is induced by the interaction of specific nutrient molecules with germinant receptors (GRs) localized in the spore's inner membrane. GRs typically consist of three subunits referred to as A, B, and C, although functions of individual subunits are not known. Here we present the crystal structure of the N-terminal domain (NTD) of the A subunit of the Bacillus megaterium GerK3 GR, revealing two distinct globular subdomains bisected by a cleft, a fold with strong homology to substrate-binding proteins in bacterial ABC transporters. Molecular docking, chemical shift perturbation measurement, and mutagenesis coupled with spore germination analyses support a proposed model that the interface between the two subdomains in the NTD of GR A subunits serves as the germinant binding site and plays a critical role in spore germination. Our findings provide a conceptual framework for understanding the germinant recruitment mechanism by which GRs trigger spore germination.


Assuntos
Bacillus megaterium/metabolismo , Proteínas de Bactérias/metabolismo , Subunidades Proteicas/metabolismo , Esporos Bacterianos/metabolismo , Simulação de Acoplamento Molecular , Domínios Proteicos
13.
Chemosphere ; 231: 184-193, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31129399

RESUMO

Hydrolyzed polyacrylamide (HPAM) is the most widely used water-soluble linear polymer with high molecular weight in polymer flooding. Microbiological degradation is an environment-friendly and effective method of treating HPAM-containing oilfield produced water. In this study, a strain SZK-5 that could degrade HPAM was isolated from soil contaminated by oilfield produced water. Based on morphological, biochemical characteristics and 16S rDNA sequence homology analysis, the strain was identified as Bacillus megaterium. The biodegradation capability of strain SZK-5 was determined by incubation in a mineral salt medium (MSM) containing HPAM under different environmental conditions, showing 55.93% of the HPAM removed after 7 d of incubation under the optimum conditions ((NH4)2SO4 = 1667.9 mg L-1, temperature = 24.05 °C and pH = 8.19). Cytochrome P450 (CYP) and urease (URE) played significant roles in biological carbon and nitrogen removal, respectively. The strain SZK-5 could resist the damages caused by oxidative stress given by crude oil and HPAM. To our knowledge, this is the first report about the biodegradation of HPAM by B. megaterium. These results suggest that strain SZK-5 might be a new auxiliary microbiological resource for the biodegradation of HPAM residue in wastewater and soil.


Assuntos
Resinas Acrílicas/metabolismo , Bacillus megaterium/metabolismo , Biodegradação Ambiental , Resinas Acrílicas/análise , Antioxidantes/metabolismo , Bacillus megaterium/enzimologia , Carbono , Hidrólise , Nitrogênio , Campos de Petróleo e Gás , Petróleo , Águas Residuárias
14.
Artigo em Inglês | MEDLINE | ID: mdl-31094278

RESUMO

The aim of the present study was to isolate microorganisms able to tolerate Ni2+ and V5+ from different sites located close to a mineral mine in Guanajuato, Mexico, and then to evaluate their ability to remove metals contained in a spent catalyst. Seventeen isolates were obtained; among them seven presented a minimum inhibitory concentration (MIC) higher than 200 mg/L of Ni2+ and V5+ each. Nickel and Vanadium removal was evaluated in 9 K liquid medium added with spent catalyst at 16% (s/v) pulp density and incubated at 30 °C, 150 rpm for 7 days. Only three isolates which were coded as PRGSd-MS-2, MNSH2-AH-3, and MNSS-AH-4 showed a significant removal at the end of treatment corresponding in mg kg-1 (or percentage metal removal) of 138 (32%), 123 (29%), and 101 (24%) for Ni, respectively; and 557 (26%), 737 (34%), and 456 (21%) mg kg-1 for V, respectively. The same isolates were capable to remove also Al, Fe, As, and Mg at different extent. Cell morphology changes were observed, in comparison to the control system at the end of biological treatment as a higher quantity of spores for MNSH2-AH-3, 2 µm cells in pairs for MNSS-AH-4, also long chain-vegetative cells having inclusions into the cell surface were observed for PRGSd-MS-2. The three isolated microorganisms were identified by sequencing of the 16S gene as Bacillus thuringiensis, Bacillus megaterium, and Bacillus sp, respectively, suggesting its potential use in the treatment of this solid industrial waste.


Assuntos
Bacillus/metabolismo , Resíduos Industriais , Metais/isolamento & purificação , Metais/farmacocinética , Purificação da Água/métodos , Bacillus/classificação , Bacillus megaterium/metabolismo , Bacillus thuringiensis/metabolismo , Reatores Biológicos/microbiologia , Catálise , Humanos , Resíduos Industriais/análise , México , Níquel/isolamento & purificação , Níquel/farmacocinética , Indústria de Petróleo e Gás/métodos , Vanádio/isolamento & purificação , Vanádio/farmacocinética , Poluentes Químicos da Água/isolamento & purificação , Poluentes Químicos da Água/farmacocinética
15.
Chemistry ; 25(26): 6533-6541, 2019 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-30820987

RESUMO

Selective chemical modification of proteins plays a pivotal role for the rational design of enzymes with novel and specific functionalities. In this study, a strategic combination of genetic and chemical engineering paves the way for systematic construction of biocatalysts by tuning the product spectrum of a levansucrase from Bacillus megaterium (Bm-LS), which typically produces small levan-like oligosaccharides. The implementation of site-directed mutagenesis followed by a tyrosine-specific modification enabled control of the product synthesis: depending on the position, the modification provoked either enrichment of short oligosaccharides (up to 800 % in some cases) or triggered the formation of high molecular weight polymer. The chemical modification can recover polymerization ability in variants with defective oligosaccharide binding motifs. Molecular dynamic (MD) simulations provided insights into the effect of modifying non-native tyrosine residues on product specificity.


Assuntos
Bacillus megaterium/enzimologia , Glicosídeo Hidrolases/química , Glicosídeo Hidrolases/genética , Hexosiltransferases/química , Hexosiltransferases/genética , Oligossacarídeos/metabolismo , Tirosina/química , Bacillus megaterium/química , Bacillus megaterium/genética , Bacillus megaterium/metabolismo , Reação de Cicloadição , Frutanos/química , Frutanos/metabolismo , Glicosídeo Hidrolases/metabolismo , Hexosiltransferases/metabolismo , Simulação de Dinâmica Molecular , Mutagênese Sítio-Dirigida , Oligossacarídeos/química , Especificidade por Substrato , Tirosina/genética , Tirosina/metabolismo
16.
Int J Biol Macromol ; 129: 728-736, 2019 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-30772411

RESUMO

Bacteria that are capable of accumulating lipids in their cells as storage compounds can also produce polyhydroxyalkanoates of high technological value, depending on the specific culture conditions. The objective of this study was to utilize crude glycerol from biodiesel (CGB) as a substrate, which is a major byproduct from biodiesel production, to produce lipophilic compounds. Bacillus megaterium INCQS 425 was cultivated and evaluated for the production of lipophilic compounds and the properties of these compounds were investigated. Cultivation of the bacteria in a medium with a C:N ratio of 0.60:1 favored the accumulation of lipids by (17.5%) comprising mainly palmitic acid (13.08%), palmitoleic (39.48%), and especially oleic acid (37.02%), which imparts good characteristics to biodiesel. Meanwhile, cultivation of the bacteria in a medium with a C:N ratio of 4:1 favored the accumulation of polyhydroxyalkanoates (PHA) (3.31gL-1) mainly comprising medium and long chain PHA. Low crystallinity (<30%) and excellent thermal properties make them suitable for processes that demand high temperatures, such as extrusion. The lipids produced in the present study had satisfactory oxidative stability for the production of quality biodiesel. The polyhydroxyalkanoates produced in the study are of low cost and have promising thermal properties that justify its technological potential, thereby configuring highly competitive bioproducts.


Assuntos
Glicerol/metabolismo , Interações Hidrofóbicas e Hidrofílicas , Bacillus megaterium/metabolismo , Biocombustíveis , Biotecnologia , Ácidos Graxos/metabolismo , Peso Molecular , Poli-Hidroxialcanoatos/química , Poli-Hidroxialcanoatos/metabolismo
17.
Nat Commun ; 10(1): 587, 2019 02 04.
Artigo em Inglês | MEDLINE | ID: mdl-30718495

RESUMO

Culture contamination, end-product toxicity, and energy efficient product recovery are long-standing bioprocess challenges. To solve these problems, we propose a high-pressure fermentation strategy, coupled with in situ extraction using the abundant and renewable solvent supercritical carbon dioxide (scCO2), which is also known for its broad microbial lethality. Towards this goal, we report the domestication and engineering of a scCO2-tolerant strain of Bacillus megaterium, previously isolated from formation waters from the McElmo Dome CO2 field, to produce branched alcohols that have potential use as biofuels. After establishing induced-expression under scCO2, isobutanol production from 2-ketoisovalerate is observed with greater than 40% yield with co-produced isopentanol. Finally, we present a process model to compare the energy required for our process to other in situ extraction methods, such as gas stripping, finding scCO2 extraction to be potentially competitive, if not superior.


Assuntos
Biocombustíveis , Dióxido de Carbono/metabolismo , Bacillus megaterium/metabolismo , Butanóis/metabolismo , Fermentação , Hemiterpenos , Cetoácidos/metabolismo , Pentanóis/metabolismo
18.
J Biotechnol ; 294: 38-48, 2019 Mar 20.
Artigo em Inglês | MEDLINE | ID: mdl-30771444

RESUMO

Over the past decades, Bacillus megaterium has gained significant interest in the biotechnological industry due to its high capacity for protein production. Although many proteins have been expressed efficiently using the optimized xylose inducible system so far, there is a considerable demand for novel promoters with varying activities, particularly for the adjustment of protein levels in multi-enzyme cascades. Genome-wide microarray analyses of the industrially important B. megaterium strain MS941 were applied to identify constitutive and growth phase dependent promoters for the expression of heterologous proteins from the early exponential to the early stationary phase of bacterial growth. Fifteen putative promoter elements were selected based on differential gene expression profiles and signal intensities of the generated microarray data. The corresponding promoter activities were evaluated in B. megaterium via ß-galactosidase screening. ß-Galactosidase expression levels ranged from 15% to 130% compared to the optimized xylose inducible promoter. Apart from these constitutive promoters we also identified and characterized novel inducible promoters, which were regulated by the addition of arabinose, galactose and the commonly used allolactose analog IPTG. The potential application of the identified promoters for biotechnologically relevant processes was demonstrated by overexpression of the cholesterol oxidase II from Brevibacterium sterolicum, thus obtaining product yields of up to 1.13 g/l/d. The provided toolbox of novel promoters offers versatile promoter strengths and will significantly contribute to harmonize protein expression in synthetic metabolic pathways, thereby pushing forward the engineering of B. megaterium as microbial cell factory for the biosynthesis and conversion of valuable compounds.


Assuntos
Bacillus megaterium/genética , Regiões Promotoras Genéticas , Bacillus megaterium/metabolismo , Colesterol Oxidase , Genoma Bacteriano , Engenharia Metabólica , Análise de Sequência com Séries de Oligonucleotídeos , Pregnenolona/metabolismo , Progesterona/metabolismo , beta-Galactosidase/metabolismo
19.
J Gen Appl Microbiol ; 65(3): 137-144, 2019 Jul 19.
Artigo em Inglês | MEDLINE | ID: mdl-30568045

RESUMO

An extracellular L-asparaginase was isolated and purified from Bacillus megaterium MG1 to apparent homogeneity. The purification procedure involved a combination of ammonium sulfate precipitation, ion-exchange chromatography, and gel filtration techniques, resulting in a purification factor of 31.52 fold with a specific activity of 215 U mg-1. The molecular mass of the purified enzyme was approximately 47 kDa on SDS-PAGE and 185 kDa on native PAGE gel as well as in gel filtration column chromatography, revealing that the enzyme was a homotetramer. The Km and Vmax values of the purified enzyme were calculated to be 2.0 ⅹ 10-4 M and 1.198 mM s-1. Maximum enzyme activity was observed over a wide range of temperature and pH values with an optimum temperature of 37°C and pH 8.5. SDS and metal ions such as Fe2+, Cu2+, Mg2+, Co2+, Mn2+, and Ca2+ decreased the enzyme activity remarkably, whereas the addition of Na+ and K+ led to an increase in activity. The insensitivity of the protein in the presence of EDTA suggested that the enzyme might not essentially be a metalloprotein. Its marked stability and activity in organic solvents and reducing agents suggest that this asparaginase is highly suitable as a biotechnological tool with industrial applications.


Assuntos
Asparaginase/isolamento & purificação , Asparaginase/metabolismo , Bacillus megaterium/enzimologia , Proteínas de Bactérias/isolamento & purificação , Proteínas de Bactérias/metabolismo , Microbiologia da Água , Asparaginase/química , Asparaginase/genética , Asparagina/metabolismo , Bacillus megaterium/classificação , Bacillus megaterium/genética , Bacillus megaterium/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Ativação Enzimática , Estabilidade Enzimática , Espaço Extracelular/metabolismo , Florestas , Concentração de Íons de Hidrogênio , Índia , Cinética , Peso Molecular , Filogenia , Especificidade por Substrato , Temperatura
20.
Chemosphere ; 216: 258-270, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30384294

RESUMO

The current study describes biological changes in Bacillus megaterium A14K cells growing in the presence of 2,3,7,8-Tetrachlorinated dibenzo-p-dioxin (TCDD), the most potent congener of dioxins. The results indicate that the metabolizing of 2,3,7,8-TCDD by BmA14K was accompanied with a novel morphological and biophysical profile typified by the growth of single cells with high levels of biosurfactant production, surface hydrophobicity and cell membrane permeability. Moreover, the TCDD-grown bacteria exhibited a specific fatty acid profile characterized by low ratios of branched/straight chain fatty acids (BCFAs/SCFAs) and saturated/unsaturated fatty acids (SFAs/USFAs) with a specific "signature" due to the presence of branched chain unsaturated fatty acids (BCUFAs). This was synchronized with a significant induction of P450BM-1, an unsaturated fatty acid-metabolizing enzyme in B. megaterium. Subsequently, the profile of oxygenated fatty acids in the TCDD-grown bacteria was typified by the presence of 5,6-epoxy derived from unsaturated C15, C16 and C17 fatty acids, that were absent in control bacteria. A net increase was also detected in both hydroxylated and epoxidized fatty acids, especially those derived from C15:0 and C16:1, respectively, suggesting a specific TCDD-induced "signature" of oxygenated fatty acids in BmA14K. Overall, this study sheds light on the use of B. megaterium A14K as a promising bioindicator/biodegrader of dioxins.


Assuntos
Bacillus megaterium/metabolismo , Sistema Enzimático do Citocromo P-450/metabolismo , Poluentes Ambientais/farmacologia , Ácidos Graxos/análise , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Dibenzodioxinas Policloradas/farmacologia , Bacillus megaterium/efeitos dos fármacos , Bacillus megaterium/crescimento & desenvolvimento , Sistema Enzimático do Citocromo P-450/efeitos dos fármacos , Ácidos Graxos/metabolismo
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