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1.
J Proteomics ; 260: 104569, 2022 May 30.
Artigo em Inglês | MEDLINE | ID: mdl-35354086

RESUMO

Anoxygenic phototrophic bacteria display phenomenal metabolic plasticity leading to distinct phenotypes. Extracellular elevated glucose levels limit photosynthesis in photosynthetic organisms; diversely, cause oxidative stress with ROS generation and "diabetic" like situation in non-photosynthetic organisms. In this study, longer incubations of externally provided glucose (22 mM) inhibited photosynthetic machinery in a phototrophic bacterium, Rubrivivax benzoatilyticus. Data analysis at three time points- exponential, early and late stationary phase, uncovered dynamic protein and metabolite abundance implying metabolic rewiring led non-cultivable state in response to glucose. Protein dynamics datum suggested that proteins related to primary metabolism down-regulated prior to those of secondary metabolism. Numerous proteins for metabolism and energy generation were highly expressed during exponential phase whereas those for membrane transport/translocation and DNA repair accumulated at early and late stationary phase respectively, suggesting a programmed knock-off of phototrophic growth mode and a switch to non-cultivable state. Overall, the omics analyses explicated the metabolic adjustment associated with glucose grown cells of R. benzoatilyticus. Further, our investigation unravelled creation of oxidative stress suggesting physiological stress (oxygen limitation) might be a key player leading to a non-cultivable state in this phototrophic organism. The study, emphasizing microbial glucose intolerance, unlocks the doorway to perceive microorganisms with new perspective. SIGNIFICANCE: Anoxygenic photosynthetic bacteria (APB), thriving under diverse habitat, exhibits magnificent metabolic flexibility. Generally, phototrophy is the preferred growth mode and energy generating route for APB. But, our analyses implicated that the glucose, under phototrophic growth conditions, triggered photobleaching in an APB member, Rubrivivax benzoatilyticus. However, retention of growth along with pigmentation under chemotrophic growth mode supports that glucose gradually knocked off the phototrophic growth mode of R. benzoatilyticus and switched to an alternate energy driving route or less energy demanding non-cultivabile state. Thus, the change in lifestyle i.e. photoheterotrophic growth instead of chemotrophic perhaps, might be the prime culprit and key player in inducing the said state of non-cultivability, akin to diabetes. The study, shedding light on the plausible regulation of cultivability, unveils the programmed regulated switching between different growth modes of the organism and illuminates the importance of glucose intolerance by microorganisms. Through this investigation, we appeal that the studies on 'glucose intolerance in microorganisms' also need due attention that will perhaps change our outlook to perceive micro-organisms in relation to their physiological life style.


Assuntos
Burkholderiales , Metaboloma , Processos Fototróficos , Burkholderiales/metabolismo , Glucose/metabolismo , Fotossíntese
2.
Environ Pollut ; 302: 119079, 2022 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-35245623

RESUMO

The excessive proliferation of Microcystis aeruginosa can lead to ecological damage, economic losses, and threaten animal and human health. For controlling Microcystis blooms, microorganism-based methods have attracted much attention from researchers because of their eco-friendliness and species-specificity. Herein, we first found that a Paucibacter strain exhibits algicidal activity against M. aeruginosa and microcystin degradation capability. The algicidal activity of DH15 (2.1 × 104 CFU/ml) against M. aeruginosa (2 × 106 cells/ml) was 94.9% within 36 h of exposure. DH15 also degraded microcystin (1.6 mg/L) up to 62.5% after 72 h. We demonstrated that the algicidal activity of DH15 against M. aeruginosa can be mediated by physical attachment and indirect attack: (1) Both washed cells and cell-free supernatant could kill M. aeruginosa efficiently; (2) Treatment with DH15 cell-free supernatants caused oxidative stress, altered the fatty acid profile, and damaged photosynthetic system, carbohydrate, and protein metabolism in M. aeruginosa. The combination of direct and indirect attacks supported that strain DH15 exerts high algicidal activity against M. aeruginosa. The expression of most key genes responsible for photosynthesis, antioxidant activity, microcystin synthesis, and other metabolic pathways in M. aeruginosa was downregulated. Strain DH15, with its microcystin degradation capacity, can overcome the trade-off between controlling Microcystis blooms and increasing microcystin concentration. Our findings suggest that strain DH15 possesses great potential to control outbreaks of Microcystis blooms.


Assuntos
Agentes de Controle Biológico , Burkholderiales , Microcystis , Agentes de Controle Biológico/metabolismo , Agentes de Controle Biológico/farmacologia , Burkholderiales/metabolismo , Herbicidas/metabolismo , Microcistinas/metabolismo , Microcystis/metabolismo , Fotossíntese
3.
J Hazard Mater ; 429: 128267, 2022 05 05.
Artigo em Inglês | MEDLINE | ID: mdl-35091192

RESUMO

Biodegradation of polyethylene terephthalate (PET) is one of fundamental ways to solve plastic pollution. As various microbial hydrolases have an extra domain unlike PETase from Ideonella sakaiensis (IsPETase), research on the role of these extra domain in PET hydrolysis is crucial for the identification and selection of a novel PET hydrolase. Here, we report that a PET hydrolase from Burkholderiales bacterium RIFCSPLOWO2_02_FULL_57_36 (BbPETase) with an additional N-terminal domain (BbPETaseAND) shows a similar hydrolysis activity toward microcrystalline PET and a higher thermal stability than IsPETase. Based on detailed structural comparisons between BbPETase and IsPETase, we generated the BbPETaseS335N/T338I/M363I/N365G variant with an enhanced PET-degrading activity and thermal stability. We further revealed that BbPETaseAND contributes to the thermal stability of the enzyme through close contact with the core domain, but the domain might hinder the adhesion of enzyme to PET substrate. We suggest that BbPETase is an enzyme in the evolution of efficient PET degradation and molecular insight into a novel PET hydrolase provides a novel strategy for the development of biodegradation of PET.


Assuntos
Burkholderiales , Hidrolases , Burkholderiales/metabolismo , Hidrolases/metabolismo , Hidrólise , Polietilenotereftalatos/química , Polietilenotereftalatos/metabolismo
4.
Chemosphere ; 286(Pt 1): 131552, 2022 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-34320440

RESUMO

Bioaugmented biotrickling filter (BTF) seeded with Piscinibacter caeni MQ-18, Pseudomonas oleovorans DT4, and activated sludge was established to investigate the treatment performance and biodegradation kinetics of the gaseous mixtures of tetrahydrofuran (THF) and methyl tert-butyl ether (MTBE). Experimental results showed an enhanced startup performance with a startup period of 9 d in bioaugmented BTF (25 d in control BTF seeded with activated sludge). The interaction parameter I2,1 of control (7.462) and bioaugmented BTF (3.267) obtained by the elimination capacity-sum kinetics with interaction parameter (EC-SKIP) model indicated that THF has a stronger inhibition of MTBE biodegradation in the control BTF than in the bioaugmented BTF. Similarly, the self-inhibition EC-SKIP model quantified the positive effects of MTBE on THF biodegradation, as well as the negative effects of THF on MTBE biodegradation and the self-inhibition of MTBE and THF. Metabolic intermediate analysis, real-time quantitative polymerase chain reaction, biofilm-biomass determination, and high-throughput sequencing revealed the possible mechanism of the enhanced treatment performance and biodegradation interactions of MTBE and THF.


Assuntos
Éteres Metílicos , Pseudomonas oleovorans , Biodegradação Ambiental , Burkholderiales , Furanos , Éteres Metílicos/análise
5.
ChemSusChem ; 15(1): e202102517, 2022 Jan 10.
Artigo em Inglês | MEDLINE | ID: mdl-34914860

RESUMO

Invited for this month's cover is the BOTTLE Consortium, featuring Gregg Beckham's laboratory from NREL and John McGeehan's laboratory from the University of Portsmouth. The cover image shows the application of poly(ethylene terephthalate) (PET) hydrolase enzymes on post-consumer waste plastic, towards the development of an enzymatic PET recycling strategy. The Full Paper itself is available at 10.1002/cssc.202101932.


Assuntos
Burkholderiales , Hidrolases , Plásticos , Polietilenotereftalatos , Reciclagem
6.
Genome Biol Evol ; 13(12)2021 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-34908127

RESUMO

Tepidimonas taiwanensis is a moderately thermophilic, Gram-negative, rod-shaped, chemoorganoheterotrophic, motile bacterium. The alkaline protease producing type strain T. taiwanensis LMG 22826T was recently reported to also be a promising producer of polyhydroxyalkanoates (PHAs)-renewable and biodegradable polymers representing an alternative to conventional plastics. Here, we present its first complete genome sequence which is also the first complete genome sequence of the whole species. The genome consists of a single 2,915,587-bp-long circular chromosome with GC content of 68.75%. Genome annotation identified 2,764 genes in total while 2,634 open reading frames belonged to protein-coding genes. Although functional annotation of the genome and division of genes into Clusters of Orthologous Groups (COGs) revealed a relatively high number of 694 genes with unknown function or unknown COG, the majority of genes were assigned a function. Most of the genes, 406 in total, were involved in energy production and conversion, and amino acid transport and metabolism. Moreover, particular key genes involved in the metabolism of PHA were identified. Knowledge of the genome in connection with the recently reported ability to produce bioplastics from the waste stream of wine production makes T. taiwanensis LMG 22826T, an ideal candidate for further genome engineering as a bacterium with high biotechnological potential.


Assuntos
Burkholderiales , Poli-Hidroxialcanoatos , Proteínas de Bactérias , Burkholderiales/genética , Endopeptidases , Poli-Hidroxialcanoatos/genética , Análise de Sequência de DNA
7.
Int J Syst Evol Microbiol ; 71(10)2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-34596504

RESUMO

A novel bacterium, strain SJAQ100T, was isolated from a freshwater aquarium and was characterized taxonomically and phylogenetically. Strain SJAQ100T was a Gram-stain-negative, aerobic, rod-shaped and non-motile bacterium. The strain grew optimally with 0 % NaCl and at 25-37 °C on Reasoner's 2A agar. Phylogenetic analysis based on the 16S rRNA gene sequences revealed that the strain SJAQ100T clustered with members of Burkholderiales incertae sedis in the order Burkholderiales, but sequence similarities to known species were less than 96.5 %. The genomic DNA G+C content of strain SJAQ100T was 71.2 mol%. Genomic comparisons of strain SJAQ100T with species in the order Burkholderiales were made using the Genome-to-Genome Distance Calculator, average nucleotide identity and average amino acid identity analyses (values indicated ≤22.1, ≤78.1, and ≤68.1 % respectively). Strain SJAQ100T contained C16 : 0 and C16 : 1 ω7c/C16 : 1 ω6c as major fatty acids and Q-8 as the major quinone. The major polyamines were putrescine and cadaverine. Strain SJAQ100T contained phosphatidylethanolamine and diphosphatidylglycerol as major polar lipids. Based on the genotypic, chemotaxonomic and phenotypic results, strain SJAQ100T represents a novel genus and species, Aquariibacter albus gen. nov., sp. nov., which belongs to order Burkholderiales and the class Betaproteobacteria. The type strain is SJAQ100T (=KCTC 72203T=CGMCC 1.18869T=MCC 4385T).


Assuntos
Burkholderiales , Ácidos Graxos , Técnicas de Tipagem Bacteriana , Composição de Bases , DNA Bacteriano/genética , Ácidos Graxos/química , Água Doce , Fosfolipídeos , Filogenia , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Ubiquinona
8.
Sheng Wu Gong Cheng Xue Bao ; 37(9): 3268-3275, 2021 Sep 25.
Artigo em Chinês | MEDLINE | ID: mdl-34622634

RESUMO

Polyethylene terephthalate (PET) is a synthetic polymer consisting of ester bond-linked terephthalate and ethylene glycol. Tremendous amounts of PET have been produced and majority of them enters terrestrial and marine environment as wastes, posing serious threats to the global ecosystems. In 2016, a PET hydrolase from a PET-assimilating bacterium Ideonalla sakaiensis was reported and termed as IsPETase. This enzyme outperforms other PET-hydrolyzing enzymes in terms of its PET hydrolytic activity at ambient temperature, thus holds a great promise for PET biodegradation. In order to improve IsPETase activity, we conducted structure-based engineering to modify the putative substrate-binding tunnel. Among the several variants to the N233 residue of IsPETase, we discovered that the substitution of N233 with alanine increases its PET hydrolytic activity, which can be further enhanced when combined with a R280A mutation. We also determined the X-ray crystal structure of the IsPETase N233A variant, which shares nearly identical fold to the WT protein, except for an open end of subsite Ⅱ. We hypothesized that the smaller side chain of N233A variant might lead to an extended subsite Ⅱ for PET binding, which subsequently increases the enzymatic activity. Thus, this study provides new clues for further structure-based engineering of PETase.


Assuntos
Burkholderiales , Hidrolases , Polietilenotereftalatos/metabolismo , Burkholderiales/enzimologia , Hidrolases/genética , Engenharia de Proteínas
9.
Phys Chem Chem Phys ; 23(39): 22451-22465, 2021 Oct 13.
Artigo em Inglês | MEDLINE | ID: mdl-34585687

RESUMO

Modulating the distribution of various states in protein ensembles through distal sites may be promising in the evolution of enzymes in desired directions. However, the prediction of distal mutation hotspots that stabilize the favoured states from a computational perspective remains challenging. Here, we presented a strategy based on molecular dynamics (MD) and Markov state models (MSM) to predict distal mutation sites. Extensive MD combined with MSM was applied to determine the principally distributed metastable states interconverting at a slow timescale. Then, molecular docking was used to classify these states into active states and inactive ones. Distal mutation hotspots were targeted based on comparing the conformational features between active and inactive states, where mutations destabilize the inactive states and show little influence on the active state. The proposed strategy was used to explore the highly dynamic MHETase, which shows a potential application in the biodegradation of poly(ethylene terephthalate) (PET). Seven principally populated interrelated metastable states were identified, and the atomistic picture of their conformational changes was unveiled. Several residues at distal positions were found to adopt more H-bond occupancies in inactive states than active states, making them potential mutation hotspots for stabilizing the favoured conformations. In addition, the detailed mechanism revealed the significance of calcium ions at a distance from the catalytic centre in reshaping the free energy landscape. This study deepens the understanding of the conformational dynamics of α/ß hydrolases containing a lid domain and advances the study of enzymatic plastic degradation.


Assuntos
Hidrolases/metabolismo , Biodegradação Ambiental , Burkholderiales/enzimologia , Hidrolases/química , Simulação de Dinâmica Molecular , Polietilenotereftalatos/química , Polietilenotereftalatos/metabolismo , Conformação Proteica
10.
Artigo em Inglês | MEDLINE | ID: mdl-34582329

RESUMO

Strain SJQ9T, an aerobic bacterium isolated from a soil sample collected in Shanghai, PR China, was characterized using a polyphasic approach. It grew optimally at pH 7.0, 30-35 °C and in the presence of 1 % (w/v) NaCl. A comparative analysis of 16S rRNA gene sequences showed that strain SJQ9T fell within the genus Aquabacterium. The closest phylogenetic relatives of strain SJQ9T were Aquabacterium citratiphilum DSM 11900T (98.6 % sequence similarity) and Aquabacterium commune DSM 11901T (96.4 %). Cells of the strain were Gram-stain-negative, motile, non-spore-forming, rod-shaped and positive for oxidase activity and negative for catalase. The chemotaxonomic properties of strain SJQ9T were consistent with those of the genus Aquabacterium: the major fatty acid was summed feature 3 (C16 : 1 ω6c and/or C16 : 1 ω7c). The isoprenoid quinone was Q-8. The major polar lipids were phosphatidylethanolamine, phosphatidylcholine, phosphatidylglycerol and diphosphatidylglycerol. The DNA G+C content was 65.7 mol%. Strain SH9T exhibited a DNA-DNA relatedness level of 34±2 % with A. citratiphilum DSM 11900T and 28±3 % with A. commune DSM 11901T. Based on the obtained data, strain SJQ9T represents a novel species of the genus Aquabacterium, for which the name Aquabacterium soli sp. nov. is proposed. The type strain is SJQ9T (=JCM 33106T=CCTCC AB 2018284T).


Assuntos
Ácidos Graxos , Solo , Técnicas de Tipagem Bacteriana , Composição de Bases , Burkholderiales , China , DNA Bacteriano/genética , Ácidos Graxos/química , Fosfolipídeos/análise , Filogenia , Piretrinas , RNA Ribossômico 16S/genética , Análise de Sequência de DNA
11.
J Hazard Mater ; 416: 126075, 2021 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-34492896

RESUMO

The development of a superb polyethylene terephthalate (PET) hydrolyzing enzyme requires an accurate understanding of the PET decomposition mechanism. However, studies on PET degrading enzymes, including the PET hydrolase from Ideonella sakaiensis (IsPETase), have not provided sufficient knowledge of the molecular mechanisms for the hardly accessible substrate. Here, we report a novel PET hydrolase from Rhizobacter gummiphilus (RgPETase), which has a hydrolyzing activity similar to IsPETase toward microcrystalline PET but distinct behavior toward low crystallinity PET film. Structural analysis of RgPETase reveals that the enzyme shares the key structural features of IsPETase for high PET hydrolysis activity but has distinguished structures at the surface-exposed regions. RgPETase shows a unique conformation of the wobbling tryptophan containing loop (WW-loop) and change of the electrostatic surface charge on the loop dramatically affects the PET-degrading activity. We further show that effect of the electrostatic surface charge to the activity varies depending on locations. This work provides valuable information underlying the uncovered PET decomposition mechanism.


Assuntos
Burkholderiales , Polietilenotereftalatos , Hidrolases
12.
Nat Commun ; 12(1): 4347, 2021 07 23.
Artigo em Inglês | MEDLINE | ID: mdl-34301933

RESUMO

Heterologous expression of biosynthetic gene clusters (BGCs) avails yield improvements and mining of natural products, but it is limited by lacking of more efficient Gram-negative chassis. The proteobacterium Schlegelella brevitalea DSM 7029 exhibits potential for heterologous BGC expression, but its cells undergo early autolysis, hindering further applications. Herein, we rationally construct DC and DT series genome-reduced S. brevitalea mutants by sequential deletions of endogenous BGCs and the nonessential genomic regions, respectively. The DC5 to DC7 mutants affect growth, while the DT series mutants show improved growth characteristics with alleviated cell autolysis. The yield improvements of six proteobacterial natural products and successful identification of chitinimides from Chitinimonas koreensis via heterologous expression in DT mutants demonstrate their superiority to wild-type DSM 7029 and two commonly used Gram-negative chassis Escherichia coli and Pseudomonas putida. Our study expands the panel of Gram-negative chassis and facilitates the discovery of natural products by heterologous expression.


Assuntos
Produtos Biológicos/metabolismo , Burkholderiales/genética , Genoma Bacteriano/genética , Família Multigênica/genética , Proteobactérias/genética , Burkholderiaceae/genética , Burkholderiaceae/metabolismo , Burkholderiales/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Regulação Bacteriana da Expressão Gênica , Engenharia Genética/métodos , Mutação , Policetídeos/metabolismo , Proteobactérias/metabolismo , Pseudomonas putida/genética , Pseudomonas putida/metabolismo
13.
Metab Eng ; 67: 250-261, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34265401

RESUMO

Poly(ethylene terephthalate) (PET) is the most abundantly consumed synthetic polyester and accordingly a major source of plastic waste. The development of chemocatalytic approaches for PET depolymerization to monomers offers new options for open-loop upcycling of PET, which can leverage biological transformations to higher-value products. To that end, here we perform four sequential metabolic engineering efforts in Pseudomonas putida KT2440 to enable the conversion of PET glycolysis products via: (i) ethylene glycol utilization by constitutive expression of native genes, (ii) terephthalate (TPA) catabolism by expression of tphA2IIA3IIBIIA1II from Comamonas and tpaK from Rhodococcus jostii, (iii) bis(2-hydroxyethyl) terephthalate (BHET) hydrolysis to TPA by expression of PETase and MHETase from Ideonella sakaiensis, and (iv) BHET conversion to a performance-advantaged bioproduct, ß-ketoadipic acid (ßKA) by deletion of pcaIJ. Using this strain, we demonstrate production of 15.1 g/L ßKA from BHET at 76% molar yield in bioreactors and conversion of catalytically depolymerized PET to ßKA. Overall, this work highlights the potential of tandem catalytic deconstruction and biological conversion as a means to upcycle waste PET.


Assuntos
Polietilenotereftalatos , Pseudomonas putida , Adipatos , Burkholderiales , Etilenos , Hidrolases , Ácidos Ftálicos , Pseudomonas putida/genética , Rhodococcus
14.
Appl Environ Microbiol ; 87(18): e0002021, 2021 08 26.
Artigo em Inglês | MEDLINE | ID: mdl-34260304

RESUMO

Poly(ethylene terephthalate) (PET) is a commonly used synthetic plastic; however, its nonbiodegradability results in a large amount of waste accumulation that has a negative impact on the environment. Recently, a PET-degrading bacterium, Ideonella sakaiensis 201-F6 strain, was isolated, and the enzymes involved in PET digestion, PET hydrolase (PETase), and mono(2-hydroxyethyl) terephthalic acid (MHET) hydrolase (MHETase) were identified. Despite the great potentials of I. sakaiensis in bioremediation and biorecycling, approaches to studying this bacterium remain limited. In this study, to enable the functional analysis of PETase and MHETase genes in vivo, we have developed a gene disruption system in I. sakaiensis. The pT18mobsacB-based disruption vector harboring directly connected 5'- and 3'-flanking regions of the target gene for homologous recombination was introduced into I. sakaiensis cells via conjugation. First, we deleted the orotidine 5'-phosphate decarboxylase gene (pyrF) from the genome of the wild-type strain, producing the ΔpyrF strain with 5-fluoroorotic acid (5-FOA) resistance. Next, using the ΔpyrF strain as a parent strain and pyrF as a counterselection marker, we disrupted the genes for PETase and MHETase. The growth of both Δpetase and Δmhetase strains on terephthalic acid (TPA; one of the PET hydrolytic products) was comparable to that of the parent strain. However, these mutant strains dramatically decreased the growth level on PET to that on a no-carbon source. Moreover, the Δpetase strain completely abolished PET degradation capacity. These results demonstrate that PETase and MHETase are essential for I. sakaiensis metabolism of PET. IMPORTANCE The poly(ethylene terephthalate) (PET)-degrading bacterium Ideonella sakaiensis possesses two unique enzymes able to serve in PET hydrolysis. PET hydrolase (PETase) hydrolyzes PET into mono(2-hydroxyethyl) terephthalic acid (MHET), and MHET hydrolase (MHETase) hydrolyzes MHET into terephthalic acid (TPA) and ethylene glycol (EG). These enzymes have attracted global attention, as they have potential to be used for bioconversion of PET. Compared to many in vitro studies, including biochemical and crystal structure analyses, few in vivo studies have been reported. Here, we developed a targeted gene disruption system in I. sakaiensis, which was then applied for constructing Δpetase and Δmhetase strains. Growth of these disruptants revealed that PETase is the sole enzyme responsible for PET degradation in I. sakaiensis, while PETase and MHETase play essential roles in its PET assimilation.


Assuntos
Proteínas de Bactérias/genética , Burkholderiales/genética , Burkholderiales/metabolismo , Hidrolases/genética , Polietilenotereftalatos/metabolismo , Proteínas de Bactérias/metabolismo , Etilenoglicol/metabolismo , Genes Bacterianos , Hidrolases/metabolismo , Hidrólise , Engenharia Metabólica , Ácidos Ftálicos/metabolismo , Reciclagem
15.
Sci Rep ; 11(1): 13745, 2021 07 02.
Artigo em Inglês | MEDLINE | ID: mdl-34215802

RESUMO

Tomato (Solanum lycopersicum L.) is an important vegetable cultivated around the world. Under field conditions, tomato can be negatively affected by water scarcity in arid and semiarid regions. The application of native plant growth-promoting rhizobacteria (PGPR) isolated from arid environments has been proposed as an inoculant to mitigate abiotic stresses in plants. In this study, we evaluated rhizobacteria from Cistanthe longiscapa (syn Calandrinia litoralis and Calandrinia longiscapa), a representative native plant of flowering desert (FD) events (Atacama Desert, Chile), to determine their ability to reduce water scarcity stress on tomato seedlings. The isolated bacterial strains were characterized with respect to their PGPR traits, including P solubilization, 1-aminocyclopropane-1-carboxylate deaminase activity, and tryptophan-induced auxin and exopolysaccharide production. Three PGPR consortia were formulated with isolated Bacillus strains and then applied to tomato seeds, and then, the seedlings were exposed to different levels of water limitations. In general, tomato seeds and seedlings inoculated with the PGPR consortia presented significantly (P ≤ 0.05) greater plant growth (48 to 60 cm of height and 171 to 214 g of weight) and recovery rates (88 to 100%) compared with those without inoculation (37 to 51 cm of height; 146 to 197 g of fresh weight; 54 to 92% of recovery) after exposure to a lack of irrigation over different time intervals (24, 72 and 120 h) before transplantation. Our results revealed the effectiveness of the formulated PGPR consortia from FD to improve the performance of inoculated seeds and seedlings subjected to water scarcity; thus, the use of these consortia can represent an alternative approach for farmers facing drought events and water scarcity associated with climate change in semiarid and arid regions worldwide.


Assuntos
Burkholderiales/metabolismo , Lycopersicon esculentum/crescimento & desenvolvimento , Desenvolvimento Vegetal , Plântula/crescimento & desenvolvimento , Burkholderiales/crescimento & desenvolvimento , Chile , Secas , Germinação/fisiologia , Ácidos Indolacéticos/metabolismo , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/microbiologia , Sementes/crescimento & desenvolvimento , Microbiologia do Solo , Insegurança Hídrica
16.
Int J Biol Macromol ; 184: 551-557, 2021 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-34171255

RESUMO

Modified potato starch with slower digestion may aid the development of new starch derivatives with improved nutritional values, and strategies to increase nutritional fractions such as resistant starch (RS) are desired. In this study, a correspondence between starch structure and enzymatic resistance was provided based on the efficient branching enzyme AqGBE, and modified starches with different amylose content (Control, 100%; PS1, 90%; PS2, 72%; PS3, 32%; PS4, 18%) were prepared. Through SEM observation, NMR and X-ray diffraction analyses, we identified that an increased proportion of α-1,6-linked branches in potato starch changes its state of granule into large pieces with crystallinity. Molecular weight and chain-length distribution analysis showed a decrease of molecular weight (from 1.1 × 106 to 1.1 × 105 g/mol) without an obvious change of chain-length distribution in PS1, while PS2-4 exhibited an increased proportion of DP 6-12 with a stable molecular weight distribution, indicating a distinct model of structural modification by AqGBE. The enhancement of peak viscosity was related to increased hydrophobic interactions and pieces state of PS1, while the contents of SDS and RS in PS1 increased by 37.7 and 49.4%, respectively. Our result provides an alternative way to increase the RS content of potato starch by branching modification.


Assuntos
Enzima Ramificadora de 1,4-alfa-Glucana/metabolismo , Burkholderiales/enzimologia , Solanum tuberosum/química , Amido/química , Amilose/química , Proteínas de Bactérias/metabolismo , Cristalografia por Raios X , Estrutura Molecular , Peso Molecular , Viscosidade , Difração de Raios X
17.
BMC Genomics ; 22(1): 464, 2021 Jun 22.
Artigo em Inglês | MEDLINE | ID: mdl-34157973

RESUMO

BACKGROUND: Cylindrospermopsin is a highly persistent cyanobacterial secondary metabolite toxic to humans and other living organisms. Strain OF001 and A210 are manganese-oxidizing bacteria (MOB) able to transform cylindrospermopsin during the oxidation of Mn2+. So far, the enzymes involved in manganese oxidation in strain OF001 and A210 are unknown. Therefore, we analyze the genomes of two cylindrospermopsin-transforming MOB, Pseudomonas sp. OF001 and Rubrivivax sp. A210, to identify enzymes that could catalyze the oxidation of Mn2+. We also investigated specific metabolic features related to pollutant degradation and explored the metabolic potential of these two MOB with respect to the role they may play in biotechnological applications and/or in the environment. RESULTS: Strain OF001 encodes two multicopper oxidases and one haem peroxidase potentially involved in Mn2+ oxidation, with a high similarity to manganese-oxidizing enzymes described for Pseudomonas putida GB-1 (80, 83 and 42% respectively). Strain A210 encodes one multicopper oxidase potentially involved in Mn2+ oxidation, with a high similarity (59%) to the manganese-oxidizing multicopper oxidase in Leptothrix discophora SS-1. Strain OF001 and A210 have genes that might confer them the ability to remove aromatic compounds via the catechol meta- and ortho-cleavage pathway, respectively. Based on the genomic content, both strains may grow over a wide range of O2 concentrations, including microaerophilic conditions, fix nitrogen, and reduce nitrate and sulfate in an assimilatory fashion. Moreover, the strain A210 encodes genes which may convey the ability to reduce nitrate in a dissimilatory manner, and fix carbon via the Calvin cycle. Both MOB encode CRISPR-Cas systems, several predicted genomic islands, and phage proteins, which likely contribute to their genome plasticity. CONCLUSIONS: The genomes of Pseudomonas sp. OF001 and Rubrivivax sp. A210 encode sequences with high similarity to already described MCOs which may catalyze manganese oxidation required for cylindrospermopsin transformation. Furthermore, the analysis of the general metabolism of two MOB strains may contribute to a better understanding of the niches of cylindrospermopsin-removing MOB in natural habitats and their implementation in biotechnological applications to treat water.


Assuntos
Alcaloides , Burkholderiales/enzimologia , Manganês , Oxirredutases , Pseudomonas/enzimologia , Burkholderiales/genética , Genoma Bacteriano , Leptothrix , Oxirredução , Oxirredutases/metabolismo , Pseudomonas/genética
18.
J Chem Inf Model ; 61(6): 3041-3051, 2021 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-34085821

RESUMO

The environmental problems derived from the generalized plastic consumption and disposal could find a friendly solution in enzymatic biodegradation. Recently, two hydrolases from Ideonella sakaiensis 201-F6 and the metagenome-derived leaf-branch compost cutinase (LCC), more specially the improved ICCG variant, have revealed degradation activity toward poly ethylene terephthalate (PET). In the present study, the reaction mechanism of this polymer breakage is studied at an atomic level by multiscale QM/MM molecular dynamics simulations, using semiempirical and DFT Hamiltonians to describe the QM region. The obtained free energy surfaces confirmed a characteristic four-step path for both systems, with activation energies in agreement with the experimental observations. Structural analysis of the evolution of the active site along the reaction progress and the study of electrostatic effects generated by the proteins reveal the similarity in the behavior of the active site of these two enzymes. The origin of the apparent better performance of the LCC-ICCG protein over PETase must be due to its capabilities of working at higher temperature and its intrinsic relationship with the crystallinity grade of the polymer. Our results may be useful for the development of more efficient enzymes in the biodegradation of PET.


Assuntos
Burkholderiales , Polietilenotereftalatos , Proteínas de Bactérias , Biodegradação Ambiental , Hidrolases
19.
Proc Natl Acad Sci U S A ; 118(23)2021 06 08.
Artigo em Inglês | MEDLINE | ID: mdl-34074785

RESUMO

Microbial interactions in aquatic environments profoundly affect global biogeochemical cycles, but the role of microparasites has been largely overlooked. Using a model pathosystem, we studied hitherto cryptic interactions between microparasitic fungi (chytrid Rhizophydiales), their diatom host Asterionella, and cell-associated and free-living bacteria. We analyzed the effect of fungal infections on microbial abundances, bacterial taxonomy, cell-to-cell carbon transfer, and cell-specific nitrate-based growth using microscopy (e.g., fluorescence in situ hybridization), 16S rRNA gene amplicon sequencing, and secondary ion mass spectrometry. Bacterial abundances were 2 to 4 times higher on individual fungal-infected diatoms compared to healthy diatoms, particularly involving Burkholderiales. Furthermore, taxonomic compositions of both diatom-associated and free-living bacteria were significantly different between noninfected and fungal-infected cocultures. The fungal microparasite, including diatom-associated sporangia and free-swimming zoospores, derived ∼100% of their carbon content from the diatom. By comparison, transfer efficiencies of photosynthetic carbon were lower to diatom-associated bacteria (67 to 98%), with a high cell-to-cell variability, and even lower to free-living bacteria (32%). Likewise, nitrate-based growth for the diatom and fungi was synchronized and faster than for diatom-associated and free-living bacteria. In a natural lacustrine system, where infection prevalence reached 54%, we calculated that 20% of the total diatom-derived photosynthetic carbon was shunted to the parasitic fungi, which can be grazed by zooplankton, thereby accelerating carbon transfer to higher trophic levels and bypassing the microbial loop. The herein termed "fungal shunt" can thus significantly modify the fate of photosynthetic carbon and the nature of phytoplankton-bacteria interactions, with implications for diverse pelagic food webs and global biogeochemical cycles.


Assuntos
Carbono/metabolismo , Quitridiomicetos/fisiologia , Diatomáceas , Cadeia Alimentar , Consórcios Microbianos , Fitoplâncton , Burkholderiales/metabolismo , Diatomáceas/metabolismo , Diatomáceas/parasitologia , Fitoplâncton/metabolismo , Fitoplâncton/parasitologia
20.
J Biotechnol ; 334: 47-50, 2021 Jun 20.
Artigo em Inglês | MEDLINE | ID: mdl-34044062

RESUMO

The large amounts of polyethylene terephthalate (PET) that enter and accumulate in the environment have posed a serious threat to global ecosystems and human health. A PET hydrolase from PET-assimilating bacterium Ideonella sakaiensis (IsPETase) that exhibits superior PET hydrolytic activity at mild conditions is attracting enormous attention in development of plastic biodegrading strategies. In order to enhance the PET hydrolysis capacity of IsPETase, we selected several polymer-binding domains that can adhere to a hydrophobic polymer surface and fused these to a previously engineered IsPETaseS121E/D186H/R280A (IsPETaseEHA) variant. We found that fusing a cellulose-binding domain (CBM) of cellobiohydrolase I from Trichoderma reesei onto the C-terminus of IsPETaseEHA showed a stimulatory effect on enzymatic hydrolysis of PET. Compared to the parental enzyme, IsPETaseEHA_CBM exhibited 71.5 % and 44.5 % higher hydrolytic activity at 30 ℃ and 40 ℃, respectively. The catalytic activity of IsPETaseEHA_CBM was increased by 86 % when the protein concentration was increased from 2.5 µg/mL to 20 µg/mL. These findings suggest that the fusion of polymer-binding module to IsPETase is a promising strategy to stimulate the enzymatic hydrolysis of PET.


Assuntos
Celulose 1,4-beta-Celobiosidase , Polietilenotereftalatos/metabolismo , Trichoderma , Burkholderiales , Celulose , Celulose 1,4-beta-Celobiosidase/genética , Ecossistema , Hidrólise , Hypocreales , Trichoderma/enzimologia
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