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1.
Food Chem ; 304: 125415, 2020 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-31479995

RESUMO

The aim of our study was to characterize the proteolytic activity of 170 Lactobacillus strains isolated from traditional Mongolian dairy products (yogurt and fermented milk), and to investigate their capacity to generate bioactive peptides during milk fermentation. All isolates were screened for proteolytic activity using skim milk agar-well diffusion test. Fifteen strains (9 Lactobacillus helveticus and 6 Lactobacillus delbrueckii subsp. bulgaricus) were then selected and further evaluated using an original strategy based on multiparametric analysis, taking into account growth rate, acidification capacity, proteolytic activity, cell envelope associated peptidase (CEP) profile and LC-MS/MS analysis of peptides. All parameters were analyzed using principal component analysis (PCA). Results showed that strain growth and acidification correlate with peptide production and that Mongolian L. helveticus strains differ from Western strains in terms of CEP distribution. The PCA revealed that CEP profiles are major determinants of ß-casein hydrolysis patterns. Strains with distinctive proteolytic activities were identified.

2.
Bioresour Technol ; 293: 122090, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31499329

RESUMO

Biofilm bioreactors have already been proven to be efficient systems for microbial lipopeptide production since they avoid foam formation. However, the cell adhesion capacities of the laboratory strain B.subtilis 168 to the biofilm bioreactor support are limited. In this work, we present a novel approach for increasing cell adhesion through the generation of filamentous and/or exopolysaccharide producing B.subtilis 168 mutants by genetic engineering. The single cell growth behavior was analyzed using time-lapse microscopy and the colonization capacities were investigated under continuous flow conditions in a drip-flow reactor. Cell adhesion could be increased three times through filamentous growth in lipopeptide producing B. subtilis 168 derivatives strains. Further restored exopolysaccharide production increased up to 50 times the cell adhesion capacities. Enhanced cell immobilization resulted in 10 times increased surfactin production. These findings will be of particular interest regarding the design of more efficient microbial cell factories for biofilm cultivation.


Assuntos
Bacillus subtilis , Lipopeptídeos , Biofilmes , Reatores Biológicos , Peptídeos Cíclicos
3.
Amino Acids ; 51(8): 1233-1240, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31197572

RESUMO

The aim of this study was to evaluate the effect of spreading the lipopeptide surfactin, for short time (10/20 s), on dentin wettability. Study groups were surfactin: 2.8; 1.4; 0.7; 0.35; and 0.175 mg/mL and a control group that received no treatment. Dentin discs (4 mm height) were prepared and polished with 600-grit SiC paper. Contact angle determinations were carried out after microbrush spreading of surfactin on dentin specimens for, respectively, 10 and 20 s. Excess liquid was removed, and after 60 s, the specimens were analyzed in a goniometer using the sessile drop method to measure the contact angle. Results were analyzed by two-way ANOVA (concentration × time) and t student, with α = 0.05. Lower contact angles were obtained for surfactin (0.7 mg/mL) spread for 10 s. However, no statistical difference was observed for surfactin (2.8 mg/mL) applied during 20 s. Higher contact angles were observed for surfactin (0.7 mg/mL) spread for 20 s. In conclusion, dentin wettability is dependent on spreading time and surfactin concentration.

4.
Front Microbiol ; 9: 2354, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30386307

RESUMO

To compensate for their amino acid auxotrophy, lactobacilli have developed the ability to hydrolyze proteins present in their environment. This proteolytic activity not only generates the free amino acids needed by the bacteria, but also a large variety of peptides, some of which are endowed with biological activities. These so-called "bioactive peptides" (BAPs) are interesting from a nutrition and healthcare perspective. The use of lactic acid bacteria (LAB) such as lactobacilli is an effective strategy for production and valorization of new BAPs. The proteolytic activity of lactobacilli is exerted in a strain- and species-dependent manner: each species exhibits different proteinase content, leading to a large variety of proteolytic activities. This underlines the high potential of Lactobacillus strains to produce novel hydrolysates and BAPs of major interest. This review aims at discussing the potential of different Lactobacillus species to release BAPs from fermentation media and processes. Strategies used for peptide production are presented. Additionally, we propose a methodology to select the most promising Lactobacillus strains as sources of BAPs. This methodology combines conventional approaches and in silico analyses.

5.
Biotechnol J ; : e1800314, 2018 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-30430761

RESUMO

Lipopeptides, such as surfactins are important biosurfactants produced by Bacillus sp. that find applications in many areas (environment, medicine, and food industries). Giving their importance, the use of simple detection methods will facilitate screening and quantification. In the present work, the authors describe a completely automated workflow for the screening of lipopeptide-producing strains, including quantification. First, isolated colonies from environmental samples are automatically picked and inoculated in 96 wells growth plate. After overnight incubation, surfactin produced in the broth is quantified, using a new sensitive fluorescent method. The method uses fluorescein (FL), which is an anionic dye at neutral to alkaline pH and forms a stable complex with the cationic surfactant cetylpiridinium chloride (CPC), quenching fluorescence. Upon addition of surfactin or other lipopeptides, fluorescein is released from the CPC-FL complex and quantified. The robustness of this method is assessed by comparing the quantification results to those conventionally measured by RP-UPLC and the results of strain screening are confirmed by MALDI-ToF analysis. The authors report for the first time the successful application of this analytical method for high-throughput screening of novel lipopeptide-producing strains.

6.
Microb Cell Fact ; 17(1): 121, 2018 Aug 04.
Artigo em Inglês | MEDLINE | ID: mdl-30077177

RESUMO

BACKGROUND: Lipopeptides are a promising group of surface-active compounds of microbial origin (biosurfactants). These diverse molecules are produced mainly by Bacillus and Pseudomonas strains. Because of their attractive physiochemical and biological properties, biosurfactants are considered to be "green and versatile molecules of the future". The main obstacles in widespread use of biosurfactants are mainly their low yields and high production costs. Pseudofactin (PF) is a lipopeptide produced by Pseudomonas fluorescens BD5. Recently, we identified two analogues, PF1 (C16-Val) and PF2 (C16-Leu), and reported that PF2 has good emulsification and foaming activities, as well as antibacterial, antifungal, anticancer, and antiadhesive properties. Reported production of PF in a mineral salt medium was approximately 10 mg/L. RESULTS: Here, we report successful high-throughput optimization of culture medium and conditions for efficient PF production using P. fluorescens BD5. Compared with production in minimal medium, PF yield increased almost 120-fold, up to 1187 ± 13.0 mg/L. Using Plackett-Burman and central composite design methodologies we identified critical factors that are important for efficient PF production, mainly high glycerol concentration, supplementation with amino acids (leucine or valine) and complex additives (e.g. tryptone), as well as high culture aeration. We also detected the shift in a ratio of produced PF analogues in response to supplementation with different amino acids. Leucine strongly induces PF2 production, while valine addition supports PF1 production. We also reported the identification of two new PF analogues: PF3 (C18-Val) and PF4 (C18-Leu). CONCLUSIONS: Identification of critical culture parameters that are important for lipopeptide production and their high yields can result in reduction of the production costs of these molecules. This may lead to the industrial-scale production of biosurfactants and their widespread use. Moreover, we produced new lipopeptide pure analogues that can be used to investigate the relationship between the structure and biological activity of lipopeptides.


Assuntos
Lipopeptídeos/metabolismo , Pseudomonas fluorescens/metabolismo , Tensoativos
7.
Environ Sci Pollut Res Int ; 25(30): 29784-29793, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-28528498

RESUMO

In this study, we have investigated the effects of three lipopeptides (fengycin, surfactin and mycosubtilin) produced by different strains of Bacillus subtilis against the phytopathogenic fungi Fusarium oxysporum f. sp. iridacearum, which affects the ornamental bulb plant populations of Iris sp. The antifungal effects were tested using minimum inhibitory concentration assay, determination of mycelium growth and spore germination inhibition rates. Also, in vivo tests on infected rhizomes and scanning electron microscopy were employed. Mycosubtilin alone and in combination with fengycin or/and surfactin showed potent inhibitory activity at concentrations as low as 5 µg ml-1 which is 100 times lower compared to Topsin M, a common chemical fungicide frequently used against fusariosis in ornamental plants. An enhancement of mycosubtilin antifungal activity was observed when it was used in combination with surfactin due to a synergistic effect. At a concentration of 20 µg ml-1, mycosubtilin inhibited the growth of the mycelium up to 49% and the spore germination ability up to 26% in comparison to control. In addition, significant changes on the macro- and micro-morphology have been observed. The antifungal activity is related to the inhibition of spore germination and the irreversible damage of the hyphae cell wall. To the best of our knowledge, this is the first attempt to propose the lipopeptides as biopesticides against the fusariosis of ornamental plants.

8.
Environ Sci Pollut Res Int ; 25(30): 29822-29833, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-28634804

RESUMO

Innovation toward ecofriendly plant protection products compatible with sustainable agriculture and healthy food is today strongly encouraged. Here, we assessed the biocontrol activity of three cyclic lipopeptides from Bacillus subtilis (mycosubtilin, M; surfactin, S; fengycin, F) and two mixtures (M + S and M + S + F) on wheat against Zymoseptoria tritici, the main pathogen on this crop. Foliar application of these biomolecules at a 100-mg L-1 concentration on the wheat cultivars Dinosor and Alixan, 2 days before fungal inoculation, provided significant reductions of disease severity. The best protection levels were recorded with the M-containing formulations (up to 82% disease reduction with M + S on Dinosor), while S and F treatments resulted in lower but significant disease reductions. In vitro and in planta investigations revealed that M-based formulations inhibit fungal growth, with half-maximal inhibitory concentrations of 1.4 mg L-1 for both M and M + S and 4.5 mg L-1 for M + S + F, thus revealing that the observed efficacy of these products may rely mainly on antifungal property. By contrast, S and F had no direct activity on the pathogen, hence suggesting that these lipopeptides act on wheat against Z. tritici as resistance inducers rather than as biofungicides. This study highlighted the efficacy of several lipopeptides from B. subtilis to biocontrol Z. tritici through likely distinct and biomolecule-dependent modes of action.

9.
Biotechnol J ; 12(7)2017 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-28636078

RESUMO

Lipopeoptides are amphiphilic compounds combining interesting physicochemical properties and biological activities. Due to their high foaming capacity in aerated bioreactor, the development of scalable bioprocesses for their production is a major bottleneck. In addition, the genes involved in the biosynthesis of these lipopeptides are mainly regulated by the quorum sensing, a global regulatory mechanism depending on cell density and known to be activated in biofilms. Several approaches have thus been considered in literature taking into account two criteria, on one side, to favor, control or avoid foam formation and on the other side, to use planktonic or immobilized (biofilm) cells. These different bioprocesses are discussed in the present review along with the purification strategies proposed for extracting and concentrating these biosurfactants.


Assuntos
Bacillus/crescimento & desenvolvimento , Reatores Biológicos/microbiologia , Lipopeptídeos/metabolismo , Bacillus/metabolismo , Biofilmes/crescimento & desenvolvimento , Células Imobilizadas/fisiologia , Fermentação , Percepção de Quorum
10.
Biotechnol J ; 12(7)2017 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-28371347

RESUMO

Surfactin, a lipopeptide produced by Bacillus subtilis, is one of the most powerful biosurfactants known. This molecule consists of a cyclic heptapeptide linked to a ß-hydroxy fatty acid chain. The isomery and the length of the fatty acid (FA) chain are responsible for the surfactin's activities. In this study, the gene codY, which encode for the global transcriptional regulator and the gene lpdV, located in the bkd operon (lpdV, bkdAA, bkdAB and bkdB genes), which is responsible for the last step of the branched chain amino acid (BCAA) degradation in acyl-CoA were deleted. The influence of these deletions on the quantitative and qualitative surfactin production was analysed. The surfactin production was quantified by RP-HPLC and the surfactin isoforms were characterized using LC-MS-MS and GC-MS analysis. The results obtained in the mutants showed an enhancement of surfactin specific production by a factor of 5.8 for the codY mutant and 1.4 for lpdV mutant. Moreover qualitative analysis of the lpdV mutant reveals that it mainly produced surfactin C14 isoform (2 fold more than the wild type) with linear FA chain. Complete analysis of the extracellular metabolites using 1 H quantitative NMR reveals a reduced production of acetoin in this mutant. This work demonstrates for the first time an original approach to overproduce specifically surfactin with C14 FA chain.


Assuntos
Bacillus subtilis/crescimento & desenvolvimento , Proteínas de Bactérias/metabolismo , Ácidos Graxos/biossíntese , Lipopeptídeos/metabolismo , Redes e Vias Metabólicas , Bacillus subtilis/genética , Proteínas de Bactérias/genética , Cromatografia Líquida , Cromatografia Gasosa-Espectrometria de Massas , Deleção de Genes , Engenharia Genética , Lipopeptídeos/genética , Óperon , Isoformas de Proteínas/metabolismo , Espectrometria de Massas em Tandem
11.
Can J Microbiol ; 63(1): 46-60, 2017 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-27912317

RESUMO

Biosurfactants are important in many areas; however, costs impede large-scale production. This work aimed to develop a global sustainable strategy for the production of biosurfactants by a novel strain of Bacillus amyloliquefaciens. Initially, Bacillus sp. strain 0G was renamed B. amyloliquefaciens subsp. plantarum (syn. Bacillus velezensis) after analysis of the gyrA and gyrB DNA sequences. Growth in modified Landy's medium produced 3 main recoverable metabolites: surfactin, fengycin, and acetoin, which promote plant growth. Cultivation was studied in the presence of renewable carbon (as glycerol) and nitrogen (as arginine) sources. While diverse kinetics of acetoin production were observed in different media, similar yields (6-8 g·L-1) were obtained after 72 h of growth. Glycerol increased surfactin-specific production, while arginine increased the yields of surfactin and fengycin and increased biomass significantly. The specific production of fengycin increased ∼10 times, possibly due to a connecting pathway involving arginine and ornithine. Adding value to crude extracts and biomass, both were shown to be useful, respectively, for the removal of p-xylene from contaminated water and for biodiesel production, yielding ∼70 mg·g-1 cells and glycerol, which could be recycled in novel media. This is the first study considering circular bioeconomy to lower the production costs of biosurfactants by valorisation of both microbial cells and their primary and secondary metabolites.


Assuntos
Bacillus amyloliquefaciens/metabolismo , Biocombustíveis/análise , Meios de Cultura/metabolismo , Poluentes Químicos da Água/metabolismo , Xilenos/metabolismo , Bacillus amyloliquefaciens/genética , Bacillus amyloliquefaciens/crescimento & desenvolvimento , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Biodegradação Ambiental , Biomassa , Glicerol/metabolismo , Lipopeptídeos/metabolismo , Peptídeos Cíclicos/metabolismo
12.
Bioprocess Biosyst Eng ; 40(2): 161-180, 2017 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-27738757

RESUMO

Innovations in novel enzyme discoveries impact upon a wide range of industries for which biocatalysis and biotransformations represent a great challenge, i.e., food industry, polymers and chemical industry. Key tools and technologies, such as bioinformatics tools to guide mutant library design, molecular biology tools to create mutants library, microfluidics/microplates, parallel miniscale bioreactors and mass spectrometry technologies to create high-throughput screening methods and experimental design tools for screening and optimization, allow to evolve the discovery, development and implementation of enzymes and whole cells in (bio)processes. These technological innovations are also accompanied by the development and implementation of clean and sustainable integrated processes to meet the growing needs of chemical, pharmaceutical, environmental and biorefinery industries. This review gives an overview of the benefits of high-throughput screening approach from the discovery and engineering of biocatalysts to cell culture for optimizing their production in integrated processes and their extraction/purification.


Assuntos
Enzimas/biossíntese , Enzimas/química , Enzimas/genética , Engenharia de Proteínas/métodos , Catálise
13.
Biosystems ; 149: 113-124, 2016 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-27769750

RESUMO

We wish to predict changes of reaction networks with partial kinetic information that lead to target changes of their steady states. The changes may be either increases or decreases of influxes, reaction knockouts, or multiple changes of these two kinds. Our prime applications are knockout prediction tasks for metabolic and regulation networks. In a first step, we propose a formal modeling language for reaction networks with partial kinetic information. The modeling language has a graphical syntax reminiscent to Petri nets. Each reaction in a model comes with a partial description of its kinetics, based on a similarity relation on kinetic functions that we introduce. Such partial descriptions are able to model the regulation of existing metabolic networks for which precise kinetic knowledge is usually not available. In a second step, we develop prediction algorithms that can be applied to any reaction network modeled in our language. These algorithms perform qualitative reasoning based on abstract interpretation, by which the kinetic unknowns are abstracted away. Given a reaction network, abstract interpretation produces a finite domain constraint in a novel class. We show how to solve these finite domain constraints with an existing finite domain constraint solver, and how to interpret the solution sets as predictions of multiple reaction knockouts that lead to a desired change of the steady states. We have implemented the prediction algorithm and integrated it into a prediction tool. This journal article extends the two conference papers John et al. (2013) and Niehren et al. (2015) while adding a new prediction algorithm for multiple gene knockouts. An application to single gene knockout prediction for surfactin overproduction was presented in Coutte et al. (2015). It illustrates the adequacy of the model-based predictions made by our algorithm in the wet lab.


Assuntos
Redes e Vias Metabólicas , Modelos Biológicos , Animais , Previsões , Humanos , Cinética , Redes e Vias Metabólicas/fisiologia
14.
Bioresour Technol ; 218: 944-52, 2016 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-27447921

RESUMO

Culture medium elements were analysed by a screening DoE to identify their influence in surfactin specific production by a surfactin constitutive overproducing Bacillus subtilis strain. Statistics pointed the major enhancement caused by high glutamic acid concentrations, as well as a minor positive influence of tryptophan and glucose. Successively, a central composite design was performed in microplate bioreactors using a BioLector®, in which variations of these impressive parameters, glucose, glutamic acid and tryptophan concentrations were selected for optimization of product-biomass yield (YP/X). Results were exploited in combination with a RSM. In absolute terms, experiments attained an YP/X 3.28-fold higher than those obtained in Landy medium, a usual culture medium used for lipopeptide production by B. subtilis. Therefore, two medium compositions for enhancing biomass and surfactin specific production were proposed and tested in continuous regime in a bubbleless membrane bioreactor. An YP/X increase of 2.26-fold was observed in bioreactor scale.


Assuntos
Bacillus subtilis , Reatores Biológicos/microbiologia , Lipopeptídeos/química , Peptídeos Cíclicos/química , Bacillus subtilis/metabolismo , Biomassa , Meios de Cultura/química
15.
Biotechnol J ; 10(8): 1216-34, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-26220295

RESUMO

A Bacillus subtilis mutant strain overexpressing surfactin biosynthetic genes was previously constructed. In order to further increase the production of this biosurfactant, our hypothesis is that the surfactin precursors, especially leucine, must be overproduced. We present a three step approach for leucine overproduction directed by methods from computational biology. Firstly, we develop a new algorithm for gene knockout prediction based on abstract interpretation, which applies to a recent modeling language for reaction networks with partial kinetic information. Secondly, we model the leucine metabolic pathway as a reaction network in this language, and apply the knockout prediction algorithm with the target of leucine overproduction. Out of the 21 reactions corresponding to potential gene knockouts, the prediction algorithm selects 12 reactions. Six knockouts were introduced in B. subtilis 168 derivatives strains to verify their effects on surfactin production. For all generated mutants, the specific surfactin production is increased from 1.6- to 20.9-fold during the exponential growth phase, depending on the medium composition. These results show the effectiveness of the knockout prediction approach based on formal models for metabolic reaction networks with partial kinetic information, and confirms our hypothesis that precursors supply is one of the main parameters to optimize surfactin overproduction.


Assuntos
Bacillus subtilis/metabolismo , Leucina/metabolismo , Lipopeptídeos/metabolismo , Modelos Biológicos , Peptídeos Cíclicos/metabolismo , Tensoativos/metabolismo , Bacillus subtilis/genética , Técnicas de Inativação de Genes , Engenharia Metabólica , Redes e Vias Metabólicas/genética
16.
Mol Plant Pathol ; 16(2): 177-87, 2015 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-25040001

RESUMO

Non-self-recognition of microorganisms partly relies on the perception of microbe-associated molecular patterns (MAMPs) and leads to the activation of an innate immune response. Bacillus subtilis produces three main families of cyclic lipopeptides (LPs), namely surfactins, iturins and fengycins. Although LPs are involved in induced systemic resistance (ISR) activation, little is known about defence responses induced by these molecules and their involvement in local resistance to fungi. Here, we showed that purified surfactin, mycosubtilin (iturin family) and plipastatin (fengycin family) are perceived by grapevine plant cells. Although surfactin and mycosubtilin stimulated grapevine innate immune responses, they differentially activated early signalling pathways and defence gene expression. By contrast, plipastatin perception by grapevine cells only resulted in early signalling activation. Gene expression analysis suggested that mycosubtilin activated salicylic acid (SA) and jasmonic acid (JA) signalling pathways, whereas surfactin mainly induced an SA-regulated response. Although mycosubtilin and plipastatin displayed direct antifungal activity, only surfactin and mycosubtilin treatments resulted in a local long-lasting enhanced tolerance to the necrotrophic fungus Botrytis cinerea in grapevine leaves. Moreover, challenge with specific strains overproducing surfactin and mycosubtilin led to a slightly enhanced stimulation of the defence response compared with the LP-non-producing strain of B. subtilis. Altogether, our results provide the first comprehensive view of the involvement of LPs from B. subtilis in grapevine plant defence and local resistance against the necrotrophic pathogen Bo. cinerea. Moreover, this work is the first to highlight the ability of mycosubtilin to trigger an immune response in plants.


Assuntos
Bacillus subtilis/metabolismo , Oligopeptídeos/farmacologia , Vitis/efeitos dos fármacos , Vitis/metabolismo , Botrytis/metabolismo , Ácidos Graxos/farmacologia , Lipopeptídeos/farmacologia , Peptídeos Cíclicos/farmacologia
17.
Appl Microbiol Biotechnol ; 98(14): 6255-64, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24723290

RESUMO

The use of surfactin and mycosubtilin as an eco-friendly alternative to control lettuce downy mildew caused by the obligate pathogen Bremia lactucae was investigated. Preliminary ecotoxicity evaluations obtained from three different tests revealed the rather low toxicity of these lipopeptides separately or in combination. The EC50 (concentration estimated to cause a 50 % response by the exposed test organisms) was about 100 mg L(-1) in Microtox assays and 6 mg L(-1) in Daphnia magna immobilization tests for mycosubtilin and 125 mg L(-1) and 25 mg L(-1) for surfactin, respectively. The toxicity of the mixture mycosubtilin/surfactin (1:1, w/w) was close to that obtained with mycosubtilin alone. In addition, the very low phytotoxic effect of these lipopeptides has been observed on germination and root growth of garden cress Lepidium sativum L. While a surfactin treatment did not influence the development of B. lactucae on lettuce plantlets, treatment with 100 mg L(-1) of mycosubtilin produced about seven times more healthy plantlets than the control samples, indicating that mycosubtilin strongly reduced the development of B. lactucae. The mixture mycosubtilin/surfactin (50:50 mg L(-1)) gave the same result on B. lactucae development as 100 mg L(-1) of mycosubtilin. The results of ecotoxicity as well as those obtained in biocontrol experiments indicated that the presence of surfactin enhances the biological activities of mycosubtilin. Mycosubtilin and surfactin were thus found to be efficient compounds against lettuce downy mildew, with low toxicity compared to the toxicity values of chemical pesticides. This is the first time that Bacillus lipopeptides have been tested in vivo against an obligate pathogen and that ecotoxic values have been given for surfactin and mycosubtilin.


Assuntos
Antifúngicos/farmacologia , Alface/microbiologia , Lipopeptídeos/farmacologia , Oomicetos/efeitos dos fármacos , Peptídeos Cíclicos/farmacologia , Controle Biológico de Vetores/métodos , Doenças das Plantas/microbiologia , Antifúngicos/toxicidade , Sinergismo Farmacológico , Lipopeptídeos/toxicidade , Lipoproteínas/farmacologia , Lipoproteínas/toxicidade , Oomicetos/crescimento & desenvolvimento , Peptídeos Cíclicos/toxicidade
18.
Bioresour Technol ; 145: 264-70, 2013 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-23583475

RESUMO

Using promoter exchange and gene knock-out strategies, two mutant strains, the so-called BBG116 and BBG125, were constructed from Bacillus subtilis wild-type strain ATCC 6633, a surfactin and mycosubtilin producer. Compared to the parental strain, both mutants overproduced constitutively mycosubtilin, while BBG125 had lost the ability to synthesize surfactin. Surprisingly, BBG125 was found to produce about 2-fold less mycosubtilin than BBG116 despite an expected higher availability of the cytoplasmic precursors and cofactors pool for biosynthesis. Further physiological characterization of BBG125 also highlighted: (i) a strong influence of temperature on mycosubtilin biosynthesis in BBG125 with a maximal productivity observed at 22°C, compared to 15 and 30°C; (ii) substantial changes in fatty acid profiles and thereby in mycosubtilin isoforms, compared to the wild-type strain; and (iii) the presence of five novel mycosubtilin isoforms. The antifungal activities of the new mix were higher than or equal to those of purified isoforms.


Assuntos
Antifúngicos , Bacillus subtilis/metabolismo , Reatores Biológicos , Biotecnologia/métodos , Lipopeptídeos/genética , Peptídeos Cíclicos/genética , Isoformas de Proteínas/biossíntese , Bacillus subtilis/genética , Cromatografia Líquida de Alta Pressão , Escherichia coli , Técnicas de Inativação de Genes , Lipoproteínas/biossíntese , Plasmídeos/genética , Especificidade da Espécie , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Espectrometria de Massas em Tandem , Temperatura Ambiente
19.
Soft Matter ; 9(40): 9572-8, 2013 Oct 28.
Artigo em Inglês | MEDLINE | ID: mdl-26029764

RESUMO

The self-assembly in aqueous solution of three lipopeptides obtained from Bacillus subtilis has been investigated. The lipopeptides surfactin, plipastatin and mycosubtilin contain distinct cyclic peptide headgroups as well as differences in alkyl chain length, branching and chain length distribution. Cryogenic transmission electron microscopy and X-ray scattering reveal that surfactin and plipastatin aggregate into 2 nm-radius spherical micelles, whereas in complete contrast mycosubtilin self-assembles into extended nanotapes based on bilayer ordering of the lipopeptides. Circular dichroism and FTIR spectroscopy indicate the presence of turn structures in the cyclic peptide headgroup. The unexpected distinct mode of self-assembly of mycosubtilin compared to the other two lipopeptides is ascribed to differences in the surfactant packing parameter. This in turn is due to specific features of the conformation of the peptide headgroup and alkyl chain branching.


Assuntos
Bacillus subtilis , Ácidos Graxos/química , Bicamadas Lipídicas/química , Lipopeptídeos/química , Micelas , Oligopeptídeos/química , Peptídeos Cíclicos/química , Microscopia Crioeletrônica , Lipoproteínas/química , Microscopia Eletrônica de Transmissão , Espectroscopia de Infravermelho com Transformada de Fourier , Difração de Raios X
20.
Appl Microbiol Biotechnol ; 87(2): 499-507, 2010 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-20221757

RESUMO

Surfactin and fengycin are lipopeptide biosurfactants produced by Bacillus subtilis. This work describes for the first time the use of bubbleless bioreactors for the production of these lipopeptides by B. subtilis ATCC 21332 with aeration by a hollow fiber membrane air-liquid contactor to prevent foam formation. Three different configurations were tested: external aeration module made from either polyethersulfone (reactor BB1) or polypropylene (reactor BB2) and a submerged module in polypropylene (reactor BB3). Bacterial growth, glucose consumption, lipopeptide production, and oxygen uptake rate were monitored during the culture in the bioreactors. For all the tested membranes, the bioreactors were of satisfactory bacterial growth and lipopeptide production. In the three configurations, surfactin production related to the culture volume was in the same range: 242, 230, and 188 mg l(-1) for BB1, BB2, and BB3, respectively. Interestingly, high differences were observed for fengycin production: 47 mg l(-1) for BB1, 207 mg l(-1) for BB2, and 393 mg l(-1) for BB3. A significant proportion of surfactin was adsorbed on the membranes and reduced the volumetric oxygen mass transfer coefficient. The degree of adsorption depended on both the material and the structure of the membrane and was higher with the submerged polypropylene membrane.


Assuntos
Bacillus subtilis/metabolismo , Reatores Biológicos/microbiologia , Microbiologia Industrial/instrumentação , Lipopeptídeos/metabolismo , Peptídeos Cíclicos/metabolismo , Tensoativos/metabolismo , Bacillus subtilis/química , Bacillus subtilis/crescimento & desenvolvimento , Biomassa , Microbiologia Industrial/métodos , Cinética , Lipopeptídeos/química , Peptídeos Cíclicos/química , Tensoativos/química
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