Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 41
Filtrar
1.
Microb Cell Fact ; 22(1): 155, 2023 Aug 16.
Artigo em Inglês | MEDLINE | ID: mdl-37582747

RESUMO

BACKGROUND: Yarrowia lipolytica is a dimorphic fungus, which switches from yeast to filament form in response to environmental conditions. For industrial purposes it is important to lock cells in the yeast or filamentous form depending on the fermentation process. yl-Hog1 kinase is a key component of the HOG signaling pathway, responsible for activating the osmotic stress response. Additionally, deletion of yl-Hog1 leads to increased filamentation in Yarrowia lipolytica, but causes significant sensitivity to osmotic stress induced by a high concentration of a carbon source. RESULTS: In this study, we tested the effect of point mutations on the function of yl-Hog1 protein kinase. The targets of modification were the phosphorylation sites (T171A-Y173A) and the active center (K49R). Introduction of the variant HOG1-49 into the hog1∆ strain partially improved growth under osmotic stress, but did not recover the yeast-like shape of the cells. The HOG1-171/173 variant was not functional, and its introduction further weakened the growth of hog1∆ strains in hyperosmotic conditions. To verify a genetic modification in filament form, we developed a new system based on green fluorescent protein (GFP) for easier screening of proper mutants. CONCLUSIONS: These results provide new insights into the functions of yl-Hog1 protein in dimorphic transition and constitute a good starting point for further genetic modification of Y. lipolytica in filament form.


Assuntos
Yarrowia , Yarrowia/metabolismo , Transdução de Sinais , Mutação , Fermentação
2.
Int J Mol Sci ; 24(7)2023 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-37047578

RESUMO

Neutrophil elastase (NE) contributes to innate antibacterial defense at both the intracellular (phagocytosis) and extracellular (degranulation, NETosis) levels. Moraxella catarrhalis, a human respiratory pathogen, can exist in an inflammatory milieu which contains NE. No data are available on the action of NE against M. catarrhalis or on the counteraction of NE-dependent host defenses by this pathogen. Using time-kill assays we found that bacteria are able to survive and replicate in the presence of NE. Transmission electron microscopy and flow cytometry studies with NE-treated bacteria revealed that while NE admittedly destabilizes the outer membrane leaflet, it does not cause cytoplasmic membrane rupture, suggesting that the enzyme does not target components that are essential for cell integrity. Using LC-MS/MS spectroscopy we determined that NE cleaved at least three virulent surface proteins in outer membrane vesicles (OMVs) of M. catarrhalis, including OMP CD, McaP, and TbpA. The cleavage of OMP CD contributes to the significant decrease in resistance to serum complement in the complement-resistant strain Mc6. The cleavage of McaP did not cause any sensitization to erythromycin nor did NE disturb its drug action. Identifying NE as a novel but subtle anti-virulence agent together with its extracellularly not-efficient bactericidal activity against M. catarrhalis may facilitate the pathogen's existence in the airways under inflammation.


Assuntos
Elastase de Leucócito , Moraxella catarrhalis , Humanos , Moraxella catarrhalis/metabolismo , Elastase de Leucócito/metabolismo , Cromatografia Líquida , Proteínas da Membrana Bacteriana Externa/metabolismo , Espectrometria de Massas em Tandem , Bactérias/metabolismo
3.
Microb Cell Fact ; 21(1): 226, 2022 Oct 28.
Artigo em Inglês | MEDLINE | ID: mdl-36307797

RESUMO

BACKGROUND: The utilization of industrial wastes as feedstock in microbial-based processes is a one of the high-potential approach for the development of sustainable, environmentally beneficial and valuable bioproduction, inter alia, lipids. Rye straw hydrolysate, a possible renewable carbon source for bioconversion, contains a large amount of xylose, inaccessible to the wild-type Yarrowia lipolytica strains. Although these oleaginous yeasts possesses all crucial genes for xylose utilization, it is necessary to induce their metabolic pathway for efficient growth on xylose and mixed sugars from agricultural wastes. Either way, biotechnological production of single cell oils (SCO) from lignocellulosic hydrolysate requires yeast genome modification or adaptation to a suboptimal environment. RESULTS: The presented Y. lipolytica strain was developed using minimal genome modification-overexpression of endogenous xylitol dehydrogenase (XDH) and xylulose kinase (XK) genes was sufficient to allow yeast to grow on xylose as a sole carbon source. Diacylglycerol acyltransferase (DGA1) expression remained stable and provided lipid overproduction. Obtained an engineered Y. lipolytica strain produced 5.51 g/L biomass and 2.19 g/L lipids from nitrogen-supplemented rye straw hydrolysate, which represents an increase of 64% and an almost 10 times higher level, respectively, compared to the wild type (WT) strain. Glucose and xylose were depleted after 120 h of fermentation. No increase in byproducts such as xylitol was observed. CONCLUSIONS: Xylose-rich rye straw hydrolysate was exploited efficiently for the benefit of production of lipids. This study indicates that it is possible to fine-tune a newly strain with as minimally genetic changes as possible by adjusting to an unfavorable environment, thus limiting multi-level genome modification. It is documented here the use of Y. lipolytica as a microbial cell factory for lipid synthesis from rye straw hydrolysate as a low-cost feedstock.


Assuntos
Yarrowia , Yarrowia/metabolismo , Biomassa , Xilose/metabolismo , Lipídeos , Carbono/metabolismo
4.
Molecules ; 26(24)2021 Dec 13.
Artigo em Inglês | MEDLINE | ID: mdl-34946639

RESUMO

The unconventional yeast Yarrowia lipolytica is used to produce erythritol from glycerol. In this study, the role of the erythrose reductase (ER) homolog YALI0B07117g in erythritol synthesis was analyzed. The deletion of the gene resulted in an increased production of mannitol (308%) and arabitol (204%) before the utilization of these polyols began. The strain overexpressing the YALI0B07117g gene was used to increase the erythritol yield from glycerol as a sole carbon source in batch cultures, resulting in a yield of 0.4 g/g. The specific consumption rate (qs) increased from 5.83 g/g/L for the WT strain to 8.49 g/g/L for the modified strain and the productivity of erythritol increased from 0.28 g/(L h) for the A101 strain to 0.41 g/(L h) for the modified strain. The application of the research may prove positive for shortening the cultivation time due to the increased rate of consumption of the substrate combined with the increased parameters of erythritol synthesis.


Assuntos
Eritritol/biossíntese , Proteínas Fúngicas , Regulação Fúngica da Expressão Gênica , Glicerol/metabolismo , Yarrowia , Eritritol/genética , Proteínas Fúngicas/biossíntese , Proteínas Fúngicas/genética , Yarrowia/genética , Yarrowia/metabolismo
5.
Arch Biochem Biophys ; 689: 108475, 2020 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-32585312

RESUMO

The unconventional yeast Yarrowia lipolytica is known as a producer of extracellular lipases. Here we overexpressed extracellular lipase (YlLip2) in yeast strain Y. lipolytica AJD ΔXΔA-Lip2 harboring the overexpression cassette of the YALI0A20350 gene under the strong hybrid promoter UAS1B16-TEF. To maintain a high level of YlLip2 production, two extracellular proteases of Y. lipolytica, AEPp and AXPp, were deleted. The purified recombinant YlLip2 presented optimal catalytic activities at 37 °C and pH 8.0. The effect of two lipopeptide biosurfactants, i.e., amphisin produced by Pseudomonas fluorescens DSS73 and viscosinamide secreted by P. fluorescens DR54, on the conformation and activity of YlLip2 was evaluated using spectral methods, surface tension, and the enzyme activity assay. YlLip2 demonstrated high tolerance of the tested biosurfactants and had greater activity retention after incubation with both biosurfactants. Finally, we observed that intrinsic fluorescence intensity of YlLip2 decreased significantly with increasing lipopeptides concentration ranging from 2.5 to 60 µM. Our results showed that both biosurfactants improve enzymatic activity of YlLip2 and might suggest better interaction of the substrate with the active site. These favorable characteristics make YlLip2 a prospective additive in the pharmaceutical, food, cosmetic, and detergent industries.


Assuntos
Lipase/metabolismo , Lipopeptídeos/metabolismo , Yarrowia/enzimologia , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Lipase/genética , Pseudomonas fluorescens/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Especificidade por Substrato , Regulação para Cima , Yarrowia/genética , Yarrowia/metabolismo
6.
Microb Cell Fact ; 19(1): 138, 2020 Jul 11.
Artigo em Inglês | MEDLINE | ID: mdl-32653007

RESUMO

BACKGROUND: During the pentose phosphate pathway (PPP), two important components, NADPH and pentoses, are provided to the cell. Previously it was shown that this metabolic pathway is a source of reducing agent for lipid synthesis from glucose in the yeast Yarrowia lipolytica. Y. lipolytica is an attractive microbial host since it is able to convert untypical feedstocks, such as glycerol, into oils, which subsequently can be transesterified to biodiesel. However, the lipogenesis process is a complex phenomenon, and it still remains unknown which genes from the PPP are involved in lipid synthesis. RESULTS: To address this problem we overexpressed five genes from this metabolic pathway: transaldolase (TAL1, YALI0F15587g), transketolase (TKL1, YALI0E06479g), ribulose-phosphate 3-epimerase (RPE1, YALI0C11880g) and two dehydrogenases, NADP+-dependent glucose-6-phosphate dehydrogenase (ZWF1, YALI0E22649g) and NADP+-dependent 6-phosphogluconate dehydrogenase (GND1, YALI0B15598g), simultaneously with diacylglycerol acyltransferase (DGA1, YALI0E32769g) and verified each resulting strain's ability to synthesize fatty acid growing on both glycerol and glucose as a carbon source. Our results showed that co-expression of DGA1 and TKL1 results in higher SCO synthesis, increasing lipid content by 40% over the control strain (DGA1 overexpression). CONCLUSIONS: Simultaneous overexpression of DGA1 and TKL1 genes results in a higher lipid titer independently from the fermentation conditions, such as carbon source, pH and YE supplementation.


Assuntos
Lipídeos/biossíntese , Transcetolase/metabolismo , Yarrowia/enzimologia , Biocombustíveis/microbiologia , Carboidratos Epimerases/genética , Carboidratos Epimerases/metabolismo , Diacilglicerol O-Aciltransferase/genética , Diacilglicerol O-Aciltransferase/metabolismo , Fermentação , Glucose/metabolismo , Glucosefosfato Desidrogenase/genética , Glucosefosfato Desidrogenase/metabolismo , Via de Pentose Fosfato , Transaldolase/genética , Transaldolase/metabolismo , Transcetolase/genética , Yarrowia/genética
7.
Microb Cell Fact ; 18(1): 176, 2019 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-31615519

RESUMO

BACKGROUND: Yarrowia lipolytica is an unconventional yeast with a huge industrial potential. Despite many advantages for biotechnological applications, it possesses enormous demand for oxygen, which is a bottleneck in large scale production. In this study a codon optimized bacterial hemoglobin from Vitreoscilla stercoraria (VHb) was overexpressed in Y. lipolytica for efficient growth and erythritol synthesis from glycerol in low-oxygen conditions. Erythritol is a natural sweetener produced by Y. lipolytica under high osmotic pressure and at low pH, and this process requires high oxygen demand. RESULTS: Under these conditions the VHb overexpressing strain showed mostly yeast-type cells resulting in 83% higher erythritol titer in shake-flask experiments. During a bioreactor study the engineered strain showed higher erythritol productivity (QERY = 0.38 g/l h) and yield (YERY = 0.37 g/g) in comparison to the control strain (QERY = 0.30 g/l h, YERY = 0.29 g/g). Moreover, low stirring during the fermentation process resulted in modest foam formation. CONCLUSIONS: This study showed that overexpression of VHb in Y. lipolytica allows for dynamic growth and efficient production of a value-added product from a low-value substrate.


Assuntos
Eritritol/biossíntese , Hemoglobinas , Microrganismos Geneticamente Modificados/metabolismo , Yarrowia , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Reatores Biológicos , Clonagem Molecular , Fermentação , Glicerol/metabolismo , Hemoglobinas/genética , Hemoglobinas/metabolismo , Engenharia Metabólica , Oxigênio/metabolismo , Vitreoscilla/metabolismo , Yarrowia/genética , Yarrowia/metabolismo
8.
Crit Rev Biotechnol ; 38(4): 620-633, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-28954540

RESUMO

Erythritol is a natural sweetener commonly used in the food and pharmaceutical industries. Produced by microorganisms as an osmoprotectant, it is an ideal sucrose substitute for diabetics or overweight persons due to its almost zero calorie content. Currently, erythritol is produced on an industrial scale through the fermentation of sugars by some yeasts, such as Moniliella sp. However, the popularity of erythritol as a sweetener is still small because of its high retail price. This creates an opportunity for further process improvement. Recent years have brought the rapid development of erythritol biosynthesis methods from the low-cost substrates, and a better understanding of the metabolic pathways leading to erythritol synthesis. The yeast Yarrowia lipolytica emerges as an organism effectively producing erythritol from pure or crude glycerol. Moreover, novel erythritol producing organisms and substrates may be taken into considerations due to metabolic engineering. This review focuses on the modification of erythritol production to use low-cost substrates and metabolic engineering of the microorganisms in order to improve yield and productivity.


Assuntos
Eritritol/biossíntese , Fermentação/fisiologia , Glicerol/metabolismo , Humanos , Engenharia Metabólica/métodos , Redes e Vias Metabólicas/fisiologia , Yarrowia/metabolismo
9.
Appl Microbiol Biotechnol ; 102(18): 7669-7678, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-29992436

RESUMO

Synthetic plastics present in everyday materials constitute the main anthropogenic debris entering the Earth's oceans. The oceans provide important and valuable resources such as food, energy, and water. They are also the main way of international trade and the main stabilizer of the climate. Hence, changes in the marine ecosystem caused by anthropogenic influences such as plastic pollution can have a dramatic impact on a global scale. Although the problem of plastics still remains unsolved, different ways are being considered to reduce their impact on the environment. One of them is to use microorganisms capable of degradation of plastic. A particularly interesting area is the application of microorganisms isolated from cold regions in view of their unique characteristics. Nevertheless, the interactions between plastic and microorganisms are still poorly known. Here, we present a review of current knowledge on plastic degradation and plastic-microorganism interactions in cold marine habitats. Moreover, we highlight the advantages of microorganisms isolated from this environment for eliminating plastic waste from ecosystems.


Assuntos
Bactérias/metabolismo , Biodegradação Ambiental , Temperatura Baixa , Ecossistema , Plásticos/metabolismo , Água do Mar/microbiologia
10.
Microb Cell Fact ; 16(1): 118, 2017 Jul 11.
Artigo em Inglês | MEDLINE | ID: mdl-28693571

RESUMO

BACKGROUND: Erythritol is a natural sweetener that is used in the food industry. It is produced as an osmoprotectant by bacteria and yeast. Due to its chemical properties, it does not change the insulin level in the blood, and therefore it can be safely used by diabetics. Previously, it has been shown that erythrose reductase (ER), which catalyzes the final step, plays a crucial role in erythritol synthesis. ER reduces erythrose to erythritol with NAD(P)H as a cofactor. Despite many studies on erythritol synthesis by Yarrowia lipolytica, the enzymes involved in this metabolic pathway have ever been described. RESULTS: The gene YALI0F18590g encoding the predicted erythrose reductase from Y. lipolytica was overexpressed, and its influence on erythritol synthesis was studied. The amino acid sequence of the Y. lipolytica ER showed a high degree of similarity to the previously described erythrose reductases from known erythritol producers, such as Candida magnoliae and Moniliella megachiliensis. Here, we found that the gene overexpression results in an enhanced titer of erythritol of 44.44 g/L (20% over the control), a yield of 0.44 g/g and productivity of 0.77 g/L/h. Moreover, on purification and characterization of the enzyme we found that it displays the highest activity at 37 °C and pH 3.0. The effects of various metal ions (Zn2+, Cu2+, Mn2+, Fe2+) on erythrose reductase were investigated. The addition of Zn2+ ions at 0.25 mM had a positive effect on the activity of erythrose reductase from Y. lipolytica, as well as on the erythritol production. CONCLUSIONS: In this study we identified, overexpressed and characterized a native erythrose reductase in Y. lipolytica. Further optimizations of this strain via metabolic pathway engineering and media optimization strategies enabled 54 g/L to be produced in a shake-flask experiment. To date, this is the first reported study employing metabolic engineering of the native gene involved in the erythritol pathway to result in a high titer of the polyol. Moreover, it indicates the importance of environmental conditions for genetic targets in metabolic engineering.


Assuntos
Aldeído Redutase/genética , Aldeído Redutase/metabolismo , Eritritol/biossíntese , Yarrowia/enzimologia , Técnicas de Cultura Celular por Lotes , Candida/enzimologia , Candida/genética , Clonagem Molecular , Eritritol/metabolismo , Glicerol/metabolismo , Concentração de Íons de Hidrogênio , Engenharia Metabólica , Redes e Vias Metabólicas , Metais/farmacologia , Proteínas Recombinantes de Fusão/metabolismo , Homologia de Sequência de Aminoácidos , Yarrowia/efeitos dos fármacos , Yarrowia/metabolismo
11.
Food Technol Biotechnol ; 55(1): 125-130, 2017 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-28559741

RESUMO

This study demonstrates the potential applicability of the UV mutant Yarrowia lipolytica MK1 for the valorisation of glycerol and erythritol production in a chemostat culture. The aim of this research is to investigate the optimal C:N ratio in the feeding medium in order to enhance erythritol production. The highest erythritol concentration, at 113.1 g/L with a volumetric erythritol production rate of 1.1 g/(L·h) and a yield of 0.57 g/g, was obtained in the feeding medium with a C:N ratio of 80:1. Moreover, no residual glycerol was observed in the culture broth during cultivation. The chemical composition of the biomass was analysed. The contents of lysine and threonine in the biomass protein amino acid profile were higher than those required by the FAO/WHO for fodder yeast.

12.
J Ind Microbiol Biotechnol ; 41(1): 57-64, 2014 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-24281394

RESUMO

Erythritol is an important natural sweetener, industrially produced only by fermentation on glucose media. Glycerol is an important renewable feedstock as it is the major by-product of the biodiesel production process; here we present an alternative way to convert this low-cost substrate into value-added products, such as erythritol. Repeated batch cultures (RBC) were performed to improve the productivity of erythritol from pure and crude glycerol. An acetate negative mutant of Yarrowia lipolytica Wratislavia K1 was found to be applicable for the production of high amounts of erythritol in RBC. When 20 % of fresh replaced medium was added, the strain Wratislavia K1 was able to produce 220 g l (-1) erythritol, which corresponded to a 0.43 g g(-1) yield and a productivity of 0.54 g l(-1) h(-1). Additionally, the activity of the culture remained stable for more than 1,000 h, i.e., 11 cycles of the repeated batch bioreactors.


Assuntos
Eritritol/biossíntese , Glicerol/metabolismo , Yarrowia/metabolismo , Técnicas de Cultura Celular por Lotes , Reatores Biológicos , Fermentação , Glucose/metabolismo
13.
Sci Total Environ ; 952: 176005, 2024 Nov 20.
Artigo em Inglês | MEDLINE | ID: mdl-39236822

RESUMO

Plastics are common synthetic materials that have been abundantly present as pollutants in natural ecosystems for the past few decades. Thus scientists have investigated the capability of plastic digestion by insects. Here we compare the effectiveness of biodegradation of the specific polymers: expanded polystyrene (EPS), polyvinyl chloride (PVC), low-density polyethylene (LDPE) and polypropylene (PP) altogether with above variants of plastics with microelements and vitamins by the mealworm - the larval form of the beetle Tenebrio molitor - and larvae of the beetle Zophobas morio, known as superworms. Z. morio beetles on all diets were able to complete their life cycle from larvae through pupae and imago, gaining 19 % and 22 % in mass on LDPE and EPS; 8 % and 7 % on PVC and PP. Mealworms (T. molitor) reared on polymers had minimal weight gain, gaining 2 % on LDPE and EPS, and a slight reduction in mass was observed when reared on PP and PVC. Not all specimens of T. molitor were able to pupate and transform to the adult stage. The results suggest that larvae of Z. morio can eat and degrade some types of plastic compounds more effectively than T. molitor. The changes in microbial gut communities were compared between these two species. The highest mass gain for Z. morio is associated with higher diversity in gut microbia and it was more diverse than that of T. molitor. Citrobacter freundii, a bacterium recognized for its ability to degrade long-chain polymers, linear hydrocarbons and cyclic hydrocarbons, was found in the microflora of Z. morio. The results confirm that superworms can survive on polymer feed. Moreover, this diet supplemented with microelements and vitamins increases the number of bacterial species and the diversity in the microbial gut.


Assuntos
Microbioma Gastrointestinal , Larva , Tenebrio , Animais , Polímeros , Besouros , Biodegradação Ambiental , Plásticos
14.
Sci Rep ; 13(1): 9129, 2023 06 05.
Artigo em Inglês | MEDLINE | ID: mdl-37277427

RESUMO

The unconventional yeast Yarrowia lipolytica produces erythritol as an osmoprotectant to adapt to osmotic stress. In this study, the array of putative erythrose reductases, responsible for the conversion of d-erythrose to erythritol, was analyzed. Single knockout and multiple knockout strains were tested for their ability to produce polyols in osmotic stress conditions. Lack of six of the reductase genes does not affect erythritol significantly, as the production of this polyol is comparable to the control strain. Deletion of eight of the homologous erythrose reductase genes resulted in a 91% decrease in erythritol synthesis, a 53% increase in mannitol synthesis, and an almost 8-fold increase in arabitol synthesis as compared to the control strain. Additionally, the utilization of glycerol was impaired in the media with induced higher osmotic pressure. The results of this research may shed new light on the production of arabitol and mannitol from glycerol by Y. lipolytica and help to develop strategies for further modification in polyol pathways in these microorganisms.


Assuntos
Yarrowia , Yarrowia/genética , Yarrowia/metabolismo , Aldeído Redutase/genética , Glicerol/metabolismo , Eritritol/metabolismo , Manitol/metabolismo
15.
Front Bioeng Biotechnol ; 10: 944228, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36061426

RESUMO

Biomass of the brown algae Fucus vesiculosus and Saccharina latissima is a promising, renewable feedstock because of the high growth rate, accessibility and content of glucose and mannitol. Saccharification of seaweeds is a simple process due to the lack of lignocellulose in the cell wall. The high content of glucose and mannitol makes these seaweeds an attractive feedstock for lipid production in the yeast Yarrowia lipolytica. This study demonstrated that hydrolysates of brown algae biomass can be applied as a substrate for synthesis of yeast biomass and lipids without any supplementation. To increase the lipid titer in yeast biomass, we employed an engineered strain of Y. lipolytica overexpressing DGA1/DGA2. In consequence, the C/N ratio has a lower impact on lipid synthesis. Moreover, the applied substrates allowed for high synthesis of unsaturated fatty acids (UFA); the level exceeded 90% in the fatty acid pool. Oleic (C18:1) and linoleic acids (C18:2) achieved the highest content. The study showed that Y. lipolytica is able to grow on the seaweed hydrolysate and produces a high content of UFA in the biomass.

16.
Sci Total Environ ; 846: 157358, 2022 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-35850328

RESUMO

There has been a growing interest in poly(ethylene terephthalate) PET degradation studies in the last few years due to its widespread use and large-scale plastic waste accumulation in the environment. One of the most promising enzymatic methods in the context of PET degradation is the use of PETase from Ideonella sakaiensis, which has been reported to be an efficient enzyme for hydrolysing ester bonds in PET. In our study, we expressed a codon-optimized PETase gene in the yeast Yarrowia lipolytica. The obtained strain was tested for its ability to degrade PET directly in culture, and a screening of different supplements that might raise the level of PET hydrolysis was performed. We also carried out long-term cultures with PET film, the surface of which was examined by scanning electron microscopy. The efficiency of PET degradation was tested based on the concentration of degradation products released, and the results showed that supplementation of the culture with olive oil resulted in 66 % higher release of terephthalic acid into the medium compared to the mutant culture without supplementation. The results indicate the possibility of ethylene glycol uptake by both strains, and, additionally, the PETase produced by the newly engineered strain hydrolyses MHET. The structure of the PET film after culture with the modified strain, meanwhile, had numerous surface defects, cracks, and deformations.


Assuntos
Polietilenotereftalatos , Yarrowia , Etilenos , Hidrolases/química , Hidrolases/genética , Hidrolases/metabolismo , Ácidos Ftálicos , Polietilenotereftalatos/química , Yarrowia/genética
17.
Sci Total Environ ; 831: 154841, 2022 Jul 20.
Artigo em Inglês | MEDLINE | ID: mdl-35358523

RESUMO

Polyethylene terephthalate (PET) is the most widely used plastic, whose global production scale causes serious problems due to it being highly non-biodegradable. The present work provides a novel approach to plastic degradation studies, which involves direct degradation of PET in the culture of a modified Y. lipolytica yeast strain extracellularly producing cutinase from Fusarium solani. In this study, we successfully accomplished a scale-up of the degradation process in culture, which is promising from the perspective of wider application of the developed method in the future. Additionally, we tested the effect of various supplements, which may increase the PET degradation efficiency in the culture of the Y. lipolytica pAD CUT_FS strain. The ability of PET decomposition was verified by the amount of the released degradation products, such as terephthalic acid (TPA) and mono-(2-hydroxyethyl)-terephthalic acid (MHET), during cultivation. We observed that the quantities of TPA and MHET released during the PET degradation process were increasing daily, and were 1.51 gL-1 and 0.45 gL-1, respectively after 240 h of the bioreactor fermentation. Analysis of the PET film by electron microscopy indicated that there was abundant damage on the surface of the material. This study also demonstrated that the engineered Y. lipolytica strain is able to degrade PET at 28 °C during fermentation. The results obtained in this study using amorphous PET powder provide a wide range of possibilities for application of the cutinase-secreting strain of Y. lipolytica on the more difficult to degrade highly crystalline PET films, PET bottles and PET melts.


Assuntos
Yarrowia , Etilenos/metabolismo , Engenharia Metabólica/métodos , Ácidos Ftálicos , Plásticos/metabolismo , Polietilenotereftalatos
18.
AMB Express ; 12(1): 12, 2022 Feb 05.
Artigo em Inglês | MEDLINE | ID: mdl-35122534

RESUMO

Cold-adapted filamentous fungal strain Geomyces sp. B10I has been reported to decompose polyesters such as poly(e-caprolactone) (PCL), poly(butylene succinate) (PBS) and poly(butylene succinate-co-butylene adipate) (PBSA). Here, we identified the enzymes of Geomyces sp. B10I, which appear to be responsible for its biodegradation activity. We compared their amino acid sequences with sequences of well-studied fungal enzymes. Partial purification of an extracellular mixture of the two enzymes, named hydrGB10I and chitGB10I, using ammonium sulfate precipitation and ionic exchange chromatography gave 14.16-fold purity. The amino acid sequence of the proteins obtained from the MALDI-TOF analysis determined the molecular mass of 77.2 kDa and 46.5 kDa, respectively. Conserved domain homology analysis revealed that both proteins belong to the class of hydrolases; hydrGB10I belongs to the glycosyl hydrolase 81 superfamily, while chitGB10I contains the domain of the glycosyl hydrolase 18 superfamily. Phylogenetic analysis suggests a distinct nature of the hydrGB10I and chitGB10I of Geomyces sp. B10I when compared with other fungal polyester-degrading enzymes described to date.

19.
Front Bioeng Biotechnol ; 9: 771133, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34917598

RESUMO

The global production of polyethylene terephthalate (PET) is estimated to reach 87.16 million metric tons by 2022. After a single use, a remarkable part of PET is accumulated in the natural environment as plastic waste. Due to high hydrophobicity and high molecular weight, PET is hardly biodegraded by wild-type microorganisms. To solve the global problem of uncontrolled pollution by PET, the degradation of plastic by genetically modified microorganisms has become a promising alternative for the plastic circular economy. In recent years many studies have been conducted to improve the microbial capacity for PET degradation. In this review, we summarize the current knowledge about metabolic engineering of microorganisms and protein engineering for increased biodegradation of PET. The focus is on mutations introduced to the enzymes of the hydrolase class-PETase, MHETase and cutinase-which in the last few years have attracted growing interest for the PET degradation processes. The modifications described in this work summarize the results obtained so far on the hydrolysis of polyethylene terephthalate based on the released degradation products of this polymer.

20.
Waste Manag ; 119: 72-81, 2021 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-33045489

RESUMO

Due to the extensive use of plastics, their quantity in the environment is constantly increasing, which creates a global problem. In the present study, we sought to isolate, test and identify Antarctic microorganisms which possess the ability to biodegrade bioplastics such as poly(ε-caprolactone) (PCL), poly(butylene succinate) (PBS) and poly(butylene succinate-co-butylene adipate) (PBSA) at low temperatures. 161 bacterial and 38 fungal isolates were isolated from 22 Antarctic soil samples. Among them, 92.16% of bacterial and 77.27% of fungal isolates formed a clear zone on emulsified PBSA, 98.04% and 81.82% on PBS and 100% and 77.27% on PCL as an additive to minimal medium at 20 °C. Based on the 16S and 18S rRNA sequences, bacterial strains were identified as species belonging to Pseudomonas and Bacillus and fungal strains as species belonging to Geomyces, Sclerotinia, Fusarium and Mortierella, while the yeast strain was identified as Hansenula anomala. In the biodegradation process conducted under laboratory conditions at 14, 20 and 28 °C, Sclerotinia sp. B11IV and Fusarium sp. B3'M strains showed the highest biodegradation activity at 20 °C (49.68% for PBSA and 33.7% for PCL, 45.99% for PBSA and 49.65% for PCL, respectively). The highest biodegradation rate for Geomyces sp. B10I was noted at 14 °C (25.67% for PBSA and 5.71% for PCL), which suggested a preference for lower temperatures (at 20 °C the biodegradation rate was only 11.34% for PBSA, and 4.46% for PCL). These data showed that microorganisms isolated from Antarctic regions are good candidates for effective plastic degradation at low temperatures.


Assuntos
Plásticos , Poliésteres , Biodegradação Ambiental , Fungos/genética , Saccharomycetales
SELEÇÃO DE REFERÊNCIAS
Detalhe da pesquisa