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1.
Microb Cell Fact ; 18(1): 163, 2019 Oct 03.
Artigo em Inglês | MEDLINE | ID: mdl-31581944

RESUMO

BACKGROUND: Sustainable production of microbial fatty acids derivatives has the potential to replace petroleum based equivalents in the chemical, cosmetic and pharmaceutical industry. Most fatty acid sources for production oleochemicals are currently plant derived. However, utilization of these crops are associated with land use change and food competition. Microbial oils could be an alternative source of fatty acids, which circumvents the issue with agricultural competition. RESULTS: In this study, we generated a chimeric microbial production system that features aspects of both prokaryotic and eukaryotic fatty acid biosynthetic pathways targeted towards the generation of long chain fatty acids. We redirected the type-II fatty acid biosynthetic pathway of Escherichia coli BL21 (DE3) strain by incorporating two homologues of the beta-ketoacyl-[acyl carrier protein] synthase I and II from the chloroplastic fatty acid biosynthetic pathway of Arabidopsis thaliana. The microbial clones harboring the heterologous pathway yielded 292 mg/g and 220 mg/g DCW for KAS I and KAS II harboring plasmids respectively. Surprisingly, beta-ketoacyl synthases KASI/II isolated from A. thaliana showed compatibility with the FAB pathway in E. coli. CONCLUSION: The efficiency of the heterologous plant enzymes supersedes the overexpression of the native enzyme in the E. coli production system, which leads to cell death in fabF overexpression and fabB deletion mutants. The utilization of our plasmid based system would allow generation of plant like fatty acids in E. coli and their subsequent chemical or enzymatic conversion to high end oleochemical products.


Assuntos
Arabidopsis/genética , Proteínas de Escherichia coli/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Ácido Graxo Sintases/metabolismo , Ácidos Graxos/biossíntese , Engenharia Metabólica , 3-Oxoacil-(Proteína de Transporte de Acila) Sintase/síntese química , 3-Oxoacil-(Proteína de Transporte de Acila) Sintase/genética , 3-Oxoacil-(Proteína de Transporte de Acila) Sintase/metabolismo , Arabidopsis/enzimologia , Proteínas de Arabidopsis/síntese química , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Vias Biossintéticas , Escherichia coli/enzimologia , Proteínas de Escherichia coli/genética , Ácido Graxo Sintases/genética , Ácidos Graxos/química , Isoenzimas/síntese química , Isoenzimas/genética , Isoenzimas/metabolismo , Plasmídeos/genética , Plasmídeos/metabolismo
2.
J Agric Food Chem ; 67(39): 10984-10993, 2019 Oct 02.
Artigo em Inglês | MEDLINE | ID: mdl-31525294

RESUMO

The objective of the present study was to reveal the effects of four types of nitrogen sources (soymeal, yeast extract, KNO3, and ammonium tartrate) on the lipid metabolism of the oleaginous fungus Mortierella alpina using untargeted lipidomics, targeted fatty acid, and reverse transcription quantitative polymerase chain reaction (RT-qPCR) analysis. Our results showed clear differences in the contents and compositions of lipids between four types of nitrogen sources. Soymeal and ammonium tartrate supplementation favored the accumulation of triglycerides with arachidonic acid (ARA) and C16-18 fatty acids, respectively. These results were further validated by our targeted fatty acid analysis. RT-qPCR analysis of related genes in M. alpina between the four nitrogen source conditions found that soymeal supplementation dramatically increased the expression of GPAT, ELOVL, and Δ12/Δ6 desaturase. Our findings provided new insights into the regulation of lipid biosynthesis in M. alpina and potential avenues for genetic manipulation and highlighted the importance of an optimal nitrogen source for ARA-rich oil production.


Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Lipídeos/biossíntese , Lipídeos/química , Espectrometria de Massas/métodos , Mortierella/metabolismo , Nitrogênio/metabolismo , Ácidos Graxos Dessaturases/genética , Ácidos Graxos Dessaturases/metabolismo , Ácidos Graxos/biossíntese , Ácidos Graxos/química , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Mortierella/química , Mortierella/enzimologia , Mortierella/genética
3.
An Acad Bras Cienc ; 91(3): e20180646, 2019 Aug 12.
Artigo em Inglês | MEDLINE | ID: mdl-31411259

RESUMO

The hepatoprotective effects of the ethanolic extracts of propolis (EEP) on alcohol-induced liver steatosis were investigated in Wistar rats. Chronic alcoholic fatty liver was induced by administration of 52% alcohol to male Wistar rats at the dose of 1% body weight for 7 weeks. Then animals were simultaneously treated with 50% ethanol solutions of EEP or normal saline at the dose of 0.1% body weight for 4 further weeks. Serological analyses and liver histopathology studies were performed to investigate the development of steatosis. Microarray analysis was conducted to investigate the alterations of hepatic gene expression profiling. Our results showed that 4-week treatment of EEP helped to restore the levels of various blood indices, liver function enzymes and the histopathology of liver tissue to normal levels. Results from the microarray analysis revealed that the hepatic expressions of genes involved in lipogenesis were significantly down-regulated by EEP treatment, while the transcriptional expressions of functional genes participating in fatty acids oxidation were markedly increased. The ability of EEP to reduce the negative effects of alcohol on liver makes propolis a potential natural product for the alternative treatment of alcoholic fatty liver.


Assuntos
Fígado Gorduroso Alcoólico/metabolismo , Hepatopatias Alcoólicas/metabolismo , Extratos Vegetais/metabolismo , Própole/metabolismo , Substâncias Protetoras/metabolismo , Alanina Transaminase/metabolismo , Animais , Apiterapia/métodos , Aspartato Aminotransferases/metabolismo , Colesterol/metabolismo , Modelos Animais de Doenças , Etanol , Ácidos Graxos/biossíntese , Fígado Gorduroso Alcoólico/tratamento farmacológico , Fígado Gorduroso Alcoólico/genética , Fígado Gorduroso Alcoólico/patologia , Hepatopatias Alcoólicas/tratamento farmacológico , Hepatopatias Alcoólicas/genética , Hepatopatias Alcoólicas/patologia , Masculino , Oxirredução , Extratos Vegetais/química , Extratos Vegetais/uso terapêutico , Própole/química , Própole/uso terapêutico , Substâncias Protetoras/química , Substâncias Protetoras/uso terapêutico , Ratos Wistar , Análise Serial de Tecidos/métodos , Transcrição Genética/genética , Triglicerídeos/metabolismo
4.
World J Microbiol Biotechnol ; 35(7): 110, 2019 Jul 06.
Artigo em Inglês | MEDLINE | ID: mdl-31280381

RESUMO

Carbon sources whether types or magnitudes were fateful in terms of stimulating growth and lipids accumulation in microalgae applied for biodiesel production. The set scenario of this work was to investigate the feasibilities of glucose (G) combining with sodium acetate (SA) carbon sources in enhancing biomass and lipid accumulation in Coccomyxa subellipsoidea. The results demonstrated that C. subellipsoidea subjected to the combination feeding of G (20 g/L) and SA (12 g/L) achieved the favorable biomass (5.22 g/L) and lipid content (52.16%). The resulting lipid productivity (388.96 mg/L/day) was 1.33- to 7.60-fold more than those of sole G or SA as well as other combinations of G and SA. Even though the total fatty acids of C. subellipsoidea cells treated with the optimal combination of G and SA showed no noticeable increment in comparison with sole G or SA, the proportion of monounsaturated C18:1 (over 48.69%) and the content of C18:3 (< 12%) were commendable in high-quality algal biodiesel production. Further, such fascinating lipid accumulation in C. subellipsoidea cells treated with G combining with SA might be attributed to that G promoted glycolysis as well as SA activated glyoxylate shunt and TCA cycle to synergistically provide sufficient acetyl-CoA precursors for lipid accumulation. These findings hinted the potential of the combination of carbon sources in enhancing the overall lipid productivity to offset alga-based biodiesel production cost and would guide other alga strains cultivation.


Assuntos
Clorófitas/crescimento & desenvolvimento , Clorófitas/metabolismo , Glucose/metabolismo , Lipídeos/biossíntese , Acetato de Sódio/metabolismo , Biocombustíveis , Biomassa , Carbono/metabolismo , Clorófitas/citologia , Meios de Cultura/química , Ácidos Graxos/biossíntese , Metabolômica , Microalgas/crescimento & desenvolvimento , Microalgas/metabolismo , Nitrogênio/metabolismo
5.
Bioresour Technol ; 291: 121783, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31326682

RESUMO

Microalgae are feedstocks for multiple product development based on algal biorefinery concept. The effects of light quality (white, red and blue light emitting diodes) and macro-element starvations on Chlorella sp. AE10 were investigated under 20% CO2 and 850 µmol m-2 d-1. Nitrogen and phosphorus starvations had negative effects on its growth rate. The biomass productivities were decreased from day 1 and the highest one was 1.90 g L-1 d-1 under white light conditions. Phosphorus starvation promoted carbohydrate accumulation under three LED light sources conditions and the highest carbohydrate content was 75.9% using red light. Blue light increased lutein content to 9.58 mg g-1. The content of saturated fatty acids was significantly increased from 37.51% under blue light and full culture medium conditions to 77.44% under blue light and nitrogen starvation conditions. Chlorella sp. AE10 was a good candidate for carbohydrate and lutein productions.


Assuntos
Carboidratos/biossíntese , Chlorella/metabolismo , Ácidos Graxos/biossíntese , Luteína/biossíntese , Biomassa , Chlorella/crescimento & desenvolvimento , Ácidos Graxos/análise , Luz , Nitrogênio/metabolismo , Fósforo/metabolismo
6.
Int J Food Microbiol ; 306: 108262, 2019 Oct 02.
Artigo em Inglês | MEDLINE | ID: mdl-31362162

RESUMO

In this study, we show that growth and prolonged storage of Listeria monocytogenes at 4 °C can promote the selection of variants with enhanced cold and heat tolerance. Enhanced cold-tolerance (ECT) variants (n = 12) were successfully isolated from a strain with impaired cold growth abilities following 84 days of storage at 4 °C in brain heart infusion broth (BHIB). Whole genome sequencing, membrane fatty acid analysis, and stress tolerance profiling were performed on the parent strain and two ECT variants: one displaying regular-sized colonies and the other displaying small colonies when grown at 37 °C on BHI agar. Under cold stress conditions, the parent strain exhibited an impaired ability to produce branched-chain fatty acids which are known to be important for cold adaptation in L.monocytogenes. The ECT variants were able to overcome this limitation, a finding which is hypothesized to be associated with the identification of two independent single-nucleotide polymorphisms in genes encoding subunits of acetyl-coA carboxylase, an enzyme critical for fatty acid biosynthesis. While the ECT phenotype was not found to be associated with improved salt (BHIB + 6% NaCl, 25 °C), acid (BHIB pH 5, 25 °C) or desiccation (33% RH, 20 °C) tolerance, the small-colony variant exhibited significantly (p < 0.05) enhanced heat tolerance at 52 °C in buffered peptone water compared to the parent strain and the other variant. The results from this study demonstrate that the continuous use of refrigeration along the food-supply chain has the potential to select for L.monocytogenes variants with enhanced cold and heat tolerance, highlighting the impact that microbial intervention strategies can have on the evolution of bacterial strains and likewise, food safety.


Assuntos
Adaptação Fisiológica/genética , Temperatura Baixa , Listeria monocytogenes/crescimento & desenvolvimento , Listeria monocytogenes/isolamento & purificação , Meios de Cultura/química , Ácidos Graxos/biossíntese , Inocuidade dos Alimentos , Listeria monocytogenes/genética , Refrigeração , Cloreto de Sódio
7.
Nat Chem Biol ; 15(7): 721-729, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31209347

RESUMO

Production of free fatty acids (FFAs) and derivatives from renewable non-food biomass by microbial fermentation is of great interest. Here, we report the development of engineered Rhodococcus opacus strains producing FFAs, fatty acid ethyl esters (FAEEs) and long-chain hydrocarbons (LCHCs). Culture conditions were optimized to produce 82.9 g l-1 of triacylglycerols from glucose, and an engineered strain with acyl-coenzyme A (CoA) synthetases deleted, overexpressing three lipases with lipase-specific foldase produced 50.2 g l-1 of FFAs. Another engineered strain with acyl-CoA dehydrogenases deleted, overexpressing lipases, foldase, acyl-CoA synthetase and heterologous aldehyde/alcohol dehydrogenase and wax ester synthase produced 21.3 g l-1 of FAEEs. A third engineered strain with acyl-CoA dehydrogenases and alkane-1 monooxygenase deleted, overexpressing lipases, foldase, acyl-CoA synthetase and heterologous acyl-CoA reductase, acyl-ACP reductase and aldehyde deformylating oxygenase produced 5.2 g l-1 of LCHCs. Metabolic engineering strategies and engineered strains developed here may help establish oleaginous biorefinery platforms for the sustainable production of chemicals and fuels.


Assuntos
Ésteres/metabolismo , Ácidos Graxos/biossíntese , Hidrocarbonetos/metabolismo , Engenharia Metabólica , Rhodococcus/metabolismo , Ésteres/química , Ácidos Graxos/química , Hidrocarbonetos/química
8.
J Agric Food Chem ; 67(25): 7005-7015, 2019 Jun 26.
Artigo em Inglês | MEDLINE | ID: mdl-31174423

RESUMO

Amino acids can enhance milk fat synthesis in bovine mammary epithelial cells (BMECs), but the molecular mechanism is not well-known. In this study, we explored the regulatory role and molecular mechanism of lysine (Lys) on milk fat synthesis induced by fatty acids (FAs). We show that Lys dose-dependently affects number of cells and milk fat synthesis, and has more stimulatory effects in the presence of FAs. Lys enhances FA-induced sterol regulatory element binding protein 1c (SREBP-1c) expression and maturation in a fatty-acid-binding protein 5 (FABP5)-dependent manner. We further show that the Lys stimulates FABP5 expression via the GPRC6A (GPCR, class C, group 6, subtype A)-PI3K (phosphatidylinositol 3-kinase) signaling. Lys dose-dependently affects GPRC6A expression and localization at the plasma membrane. In summary, our data reveals that Lys enhances FAs-stimulated SREBP-1c expression and maturation leading to milk fat synthesis via the GPRC6A-PI3K-FABP5 signaling in BMECs.


Assuntos
Bovinos/metabolismo , Células Epiteliais/metabolismo , Proteínas de Ligação a Ácido Graxo/metabolismo , Ácidos Graxos/biossíntese , Glândulas Mamárias Animais/metabolismo , Leite/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Receptores Acoplados a Proteínas-G/metabolismo , Animais , Bovinos/genética , Gorduras/metabolismo , Proteínas de Ligação a Ácido Graxo/genética , Feminino , Lisina , Glândulas Mamárias Animais/citologia , Fosfatidilinositol 3-Quinases/genética , Receptores Acoplados a Proteínas-G/genética , Transdução de Sinais , Proteína de Ligação a Elemento Regulador de Esterol 1/genética , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismo
9.
Am J Chin Med ; 47(4): 787-801, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31091973

RESUMO

American ginseng and Asian ginseng, which occupy prominent positions in the list of best-selling natural products in the West and East, are suitable for different indications in the traditional pharmacological uses. Currently, the effects of American ginseng and Asian ginseng in the protection against metabolic dysfunction and the differences between them are still unknown. Herein, an untargeted metabolomics based on liquid chromatography-quadrupole time-of-flight mass spectrometry (LC-Q-TOF-MS) was determined. The serum metabolomics and dynamic feces metabolomics revealed significant metabolic distinction between American ginseng and Asian ginseng in diet-induced obese (DIO) mice. The results show that American ginseng and Asian ginseng alleviate glucose and lipid metabolism disorder in DIO mice. A total of 45 differential metabolites were confirmed between the drug-naïve and American ginseng group, and 32 metabolites were confirmed between the drug-naïve and Asian ginseng group. Metabolic pathways analysis shows that these two ginsengs treatment dynamic rectifies metabolic disorder in DIO mice mainly via regulating linoleic acids metabolism, cysteine and methionine metabolism and biosynthesis of unsaturated fatty acid. Moreover, American ginseng's specific function in monitoring the carnitines and taurine/hypotaurine metabolism might make it more effective in meliorating lipids metabolism disorder than Asian ginseng.


Assuntos
Dieta Hiperlipídica/efeitos adversos , Glucose/metabolismo , Metabolismo dos Lipídeos/efeitos dos fármacos , Metabolômica/métodos , Obesidade/etiologia , Obesidade/metabolismo , Panax/química , Panax/classificação , Extratos Vegetais/farmacologia , Animais , Carnitina/metabolismo , Cromatografia Líquida , Cisteína/metabolismo , Ácidos Graxos/biossíntese , Ácido Linoleico/metabolismo , Masculino , Espectrometria de Massas , Metionina/metabolismo , Camundongos Endogâmicos C57BL , Obesidade/tratamento farmacológico , Fitoterapia , Extratos Vegetais/uso terapêutico , Taurina/metabolismo
10.
J Oleo Sci ; 68(6): 541-549, 2019 Jun 06.
Artigo em Inglês | MEDLINE | ID: mdl-31092798

RESUMO

Thraustochytrids, a group of marine protists, are continuously gaining attention due to their capability in producing lipids for various biotechnological applications towards foods, medicines, chemicals, and biofuels. Although various substrates, predominantly glucose, have been used as carbon source for this microalga, it is desirable to adopt cheaper and more diversified substrate to expand their application range. In this study, we aimed to examine the ability of acetate, which can be easily generated from various resources by acetogenic microorganisms, as a substrate of Aurantiochytrium limacinum SR21. As a result of flask-scale analysis, specific growth rates (µ) of the strain SR21 grown in 3% acetate- or glucose-based medium were 0.55 and 0.98 h-1, respectively. The maximum yield of total fatty acid in acetate medium was 4.8 g/L at 48 h while that in glucose medium was 6.8 g/L at 30 h, indicating that acetate has potential as substrate. Metabolome analysis was performed to comprehensively elucidate characteristic metabolic fluctuations caused by acetate assimilation and identify targets to improve the fatty acid productivity from acetate. It was found that the use of glyoxylate cycle, which bypasses release of energy molecules such as NADH and GTP, and the inhibition of utilization of compounds from TCA cycle for anabolic reactions, may cause the slow growth in acetate which has an effect also in lipid productivity. The activity of the pentose phosphate pathway was found to be weak in acetate cultivation, thus NADPH was mainly produced in malate-pyruvate cycle. Lastly, mevalonate pathway was found to be activated in acetate cultivation which additionally competes with acetyl-CoA as starting material of fatty acid synthesis.


Assuntos
Acetatos/metabolismo , Meios de Cultura , Ácidos Graxos/biossíntese , Fermentação/fisiologia , Metabolismo dos Lipídeos/fisiologia , Estramenópilas/metabolismo , Acetilcoenzima A/metabolismo , Meios de Cultura/química , Ácido Glucárico/metabolismo , Ácido Mevalônico/metabolismo , NADP/biossíntese , Via de Pentose Fosfato , Estramenópilas/crescimento & desenvolvimento
11.
Mol Cell ; 74(3): 598-608.e6, 2019 05 02.
Artigo em Inglês | MEDLINE | ID: mdl-31051140

RESUMO

RNA flow between organisms has been documented within and among different kingdoms of life. Recently, we demonstrated horizontal RNA transfer between honeybees involving secretion and ingestion of worker and royal jellies. However, how the jelly facilitates transfer of RNA is still unknown. Here, we show that worker and royal jellies harbor robust RNA-binding activity. We report that a highly abundant jelly component, major royal jelly protein 3 (MRJP-3), acts as an extracellular non-sequence-specific RNA-aggregating factor. Multivalent RNA binding stimulates higher-order assembly of MRJP-3 into extracellular ribonucleoprotein granules that protect RNA from degradation and enhance RNA bioavailability. These findings reveal that honeybees have evolved a secreted dietary RNA-binding factor to concentrate, stabilize, and share RNA among individuals. Our work identifies high-order ribonucleoprotein assemblies with functions outside cells and organisms.


Assuntos
Abelhas/genética , Ácidos Graxos/genética , Transferência Genética Horizontal/genética , Glicoproteínas/genética , Proteínas de Insetos/genética , Animais , Ácidos Graxos/biossíntese , Transição de Fase , RNA/genética , Transporte de RNA/genética , Proteínas de Ligação a RNA/genética
12.
Oxid Med Cell Longev ; 2019: 9214209, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31049141

RESUMO

The experiment was designed to clarify the effect and molecular mechanism of maternal genistein (GEN) on the lipid metabolism and developmental growth of offspring chicks. Laying broiler breeder (LBB) hens were supplemented with 40 mg/kg genistein (GEN), while the control group was fed with the low-soybean meal diet. The offspring chicks were grouped according to the mother generation with 8 replicates each. Hepatic transcriptome data revealed 3915 differentially expressed genes (DEGs, P adjusted < 0.05, fold change > 1.5 or fold change < 0.67) between chicks in the two groups. Maternal GEN activated the GH-IGF1-PI3K/Akt signaling pathway, which promoted the developmental processes and cellular amino acid metabolic processes, as well as inhibited the apoptotic process. GEN treatment significantly increased the weight gain, breast muscle percentage, and liver index in chicks. PANTHER clustering analysis suggested that maternal GEN enhanced the antioxidant activity of chicks by the upregulation of gene (SOD3, MT1, and MT4) expression. Accordingly, the activities of T-AOC and T-SOD in the liver were increased after GEN treatment. The overrepresentation tests revealed that maternal GEN influenced the glycolysis, unsaturated fatty acid biosynthesis, acyl-coenzyme A metabolism, lipid transport, and cholesterol metabolism in the chick livers. Hepatic cholesterol and long-chain fatty acid were significantly decreased after GEN treatment. However, the level of arachidonic acid was higher in the livers of the GEN-treated group compared with the CON group. Moreover, GEN treatment enhanced fatty acid ß-oxidation and upregulated PPARδ expression in the chick liver. ChIP-qPCR analysis indicated that maternal GEN might induce histone H3-K36 trimethylation in the promoter region of PPARδ gene (PPARD) through Iws1, methyltransferases. It also induced histone H4-K12 acetylation at the PPARD promoter through MYST2, which activated the PPAR signaling pathways in the chick livers. In summary, supplementing LBB hens with GEN can alter lipid metabolism in the offspring chicks through epigenetic modification and improve the antioxidative capability as well as growth performance.


Assuntos
Suplementos Nutricionais , Epigênese Genética/efeitos dos fármacos , Ácidos Graxos/biossíntese , Genisteína/farmacologia , Metabolismo dos Lipídeos/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Galinhas , Feminino , Masculino
13.
Microb Cell Fact ; 18(1): 92, 2019 May 28.
Artigo em Inglês | MEDLINE | ID: mdl-31138218

RESUMO

BACKGROUND: The carboxylate platform is a promising technology for substituting petrochemicals in the provision of specific platform chemicals and liquid fuels. It includes the chain elongation process that exploits reverse ß-oxidation to elongate short-chain fatty acids and forms the more valuable medium-chain variants. The pH value influences this process through multiple mechanisms and is central to effective product formation. Its influence on the microbiome dynamics was investigated during anaerobic fermentation of maize silage by combining flow cytometric short interval monitoring, cell sorting and 16S rRNA gene amplicon sequencing. RESULTS: Caproate and caprylate titres of up to 6.12 g L-1 and 1.83 g L-1, respectively, were achieved in a continuous stirred-tank reactor operated for 241 days. Caproate production was optimal at pH 5.5 and connected to lactate-based chain elongation, while caprylate production was optimal at pH 6.25 and linked to ethanol utilisation. Flow cytometry recorded 31 sub-communities with cell abundances varying over 89 time points. It revealed a highly dynamic community, whereas the sequencing analysis displayed a mostly unchanged core community. Eight key sub-communities were linked to caproate or caprylate production (rS > | ± 0.7|). Amongst other insights, sorting and subsequently sequencing these sub-communities revealed the central role of Bifidobacterium and Olsenella, two genera of lactic acid bacteria that drove chain elongation by providing additional lactate, serving as electron donor. CONCLUSIONS: High-titre medium-chain fatty acid production in a well-established reactor design is possible using complex substrate without the addition of external electron donors. This will greatly ease scaling and profitable implementation of the process. The pH value influenced the substrate utilisation and product spectrum by shaping the microbial community. Flow cytometric single cell analysis enabled fast, short interval analysis of this community and was coupled with 16S rRNA gene amplicon sequencing to reveal the major role of lactate-producing bacteria.


Assuntos
Ácidos Acíclicos/metabolismo , Reatores Biológicos , Ácidos Graxos/biossíntese , Ácido Láctico/metabolismo , Microbiota , Fermentação , Microbiota/genética , Microbiota/fisiologia , RNA Ribossômico 16S , Análise de Célula Única
14.
Genome Biol Evol ; 11(6): 1541-1551, 2019 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-31076758

RESUMO

The birth-and-death evolutionary model proposes that some members of a multigene family are phylogenetically stable and persist as a single copy over time, whereas other members are phylogenetically unstable and undergo frequent duplication and loss. Functional studies suggest that stable genes are likely to encode essential functions, whereas rapidly evolving genes reflect phenotypic differences in traits that diverge rapidly among species. One such class of rapidly diverging traits are insect cuticular hydrocarbons (CHCs), which play dual roles in chemical communications as short-range recognition pheromones as well as protecting the insect from desiccation. Insect CHCs diverge rapidly between related species leading to ecological adaptation and/or reproductive isolation. Because the CHC and essential fatty acid biosynthetic pathways share common genes, we hypothesized that genes involved in the synthesis of CHCs would be evolutionary unstable, whereas those involved in fatty acid-associated essential functions would be evolutionary stable. To test this hypothesis, we investigated the evolutionary history of the fatty acyl-CoA reductases (FARs) gene family that encodes enzymes in CHC synthesis. We compiled a unique data set of 200 FAR proteins across 12 Drosophila species. We uncovered a broad diversity in FAR content which is generated by gene duplications, subsequent gene losses, and alternative splicing. We also show that FARs expressed in oenocytes and presumably involved in CHC synthesis are more unstable than FARs from other tissues. Taken together, our study provides empirical evidence that a comparative approach investigating the birth-and-death evolution of gene families can identify candidate genes involved in rapidly diverging traits between species.


Assuntos
Aldeído Oxirredutases/genética , Drosophila/enzimologia , Drosophila/genética , Evolução Molecular , Animais , Drosophila/classificação , Drosophila/crescimento & desenvolvimento , Drosophila melanogaster/enzimologia , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Embrião não Mamífero/enzimologia , Ácidos Graxos/biossíntese , Duplicação Gênica , Redes e Vias Metabólicas , Filogenia
15.
Chem Pharm Bull (Tokyo) ; 67(4): 327-332, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30930436

RESUMO

Δ9-Fatty acid desaturase (Δ9-desaturase) is a rate-limiting enzyme of unsaturated fatty acid biosynthesis in animal cells and specifically introduces a cis-double bond at the Δ9-position of acyl-CoA. Since the chemical structure of fatty acids determines the physicochemical properties of cellular membrane and modulates a broad range of cellular functions, double bond introduction into a fatty acid by Δ9-desaturase should be specifically carried out. Reported crystal structures of stearoyl-CoA desaturase (SCD)1, one of the most studied Δ9-desaturases, have revealed the mechanism underlying the determination of substrate preference, as well as the position (Δ9) and conformation (cis) of double bond introduction. The crystal structures of SCD1 have also provided insights into the function of other Δ9-desaturases, including Drosophila homologs. Moreover, the amino-terminal sequences of Δ9-desaturases are shown to have unique roles in protein degradation. In this review, we introduce recent advances in the understanding of the function and regulation of Δ9-desaturase from the standpoint of protein structure.


Assuntos
Ácidos Graxos Dessaturases/química , Sequência de Aminoácidos , Animais , Ácidos Graxos Dessaturases/metabolismo , Ácidos Graxos/biossíntese , Humanos , Estrutura Terciária de Proteína , Alinhamento de Sequência , Estearoil-CoA Dessaturase/química , Estearoil-CoA Dessaturase/metabolismo , Especificidade por Substrato
16.
J Agric Food Chem ; 67(16): 4545-4552, 2019 Apr 24.
Artigo em Inglês | MEDLINE | ID: mdl-30929440

RESUMO

Long-chain hydroxy fatty acids (HFAs) are rare in nature but have many promising industrial applications. In this study, we developed a biosynthesis method to produce long-chain ω-hydroxy fatty acids. Through disruption of the acyl-CoA synthetases FAA1 and FAA4 and the fatty acyl-CoA oxidase POX1, a Saccharomyces cerevisiae strain was engineered to accumulate free fatty acids (FFAs). Subsequently, the cytochrome P450 monooxygenase CYP52M1 from Starmerella bombicola was introduced to convert FFAs to HFAs, leading to the production of C16 and C18 HFAs at the ω or ω-1 positions. Next, CYP52M1 was reconstituted with the homologous reductase S. bombicola CPR and the heterologous reductase Arabidopsis thaliana cytochrome P450 reductase. The results showed that the CYP52M1-AtCPR1 system significantly increased the hydroxylation in FFA. Moreover, a self-sufficient P450 enzyme system was constructed to achieve higher transformation efficiency. Finally, fed-batch fermentation yielded as much as 347 ± 9.2 mg/L ω-HFAs.


Assuntos
Ácidos Graxos/biossíntese , Ácidos Graxos/química , Engenharia Metabólica , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Coenzima A Ligases/genética , Coenzima A Ligases/metabolismo , Sistema Enzimático do Citocromo P-450/genética , Sistema Enzimático do Citocromo P-450/metabolismo , Fermentação , Hidroxilação , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo
17.
Biochim Biophys Acta Gene Regul Mech ; 1862(6): 643-656, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30959128

RESUMO

Gluconeogenesis is essential for blood glucose homeostasis during fasting and is regulated by various enzymes, which are encoded by gluconeogenic genes. Those genes are controlled by various transcription factors. Zinc finger and BTB domain-containing 7c (Zbtb7c, also called Kr-pok) is a BTB-POZ family transcription factor with proto-oncogenic activity. Previous findings have indicated that Zbtb7c is involved in the regulation of fatty acid biosynthesis, suggesting an involvement also in primary metabolism. We found here that fasting induced Zbtb7c expression in the mouse liver and in primary liver hepatocytes. We also observed that Zbtb7c-knockout mice have decreased blood glucose levels, so we investigated whether Zbtb7c plays a role in gluconeogenesis. Indeed, differential gene expression analysis of Zbtb7c-knockout versus wild type mouse livers showed downregulated transcription of gluconeogenic genes encoding the glucose 6-phosphatase catalytic subunit (G6pc) and phosphoenolpyruvate carboxykinase 1 (Pck1), while Zbtb7c expression upregulated these two genes, under fasting conditions. Mechanistically, we found that when complexed with histone deacetylase 3 (Hdac3), Zbtb7c binds insulin response elements (IREs) within the G6pc and Pck1 promoters. Moreover, complexed Zbtb7c deacetylated forkhead box O1 (Foxo1), thereby increasing Foxo1 binding to the G6pc and Pck1 IREs, resulting in their transcriptional activation. These results demonstrate Zbtb7c to be a crucial metabolic regulator of blood glucose homeostasis, during mammalian fasting.


Assuntos
Jejum , Regulação da Expressão Gênica , Gluconeogênese/fisiologia , Glucose-6-Fosfatase/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Fosfoenolpiruvato Carboxiquinase (GTP)/genética , Proteínas/metabolismo , Fatores de Transcrição/metabolismo , Dedos de Zinco/fisiologia , Animais , Glicemia , Proteínas de Ligação a DNA/metabolismo , Ácidos Graxos/biossíntese , Proteína Forkhead Box O1/metabolismo , Gluconeogênese/genética , Glucose/metabolismo , Glucose-6-Fosfatase/metabolismo , Células HEK293 , Células Hep G2 , Hepatócitos/metabolismo , Histona Desacetilases/metabolismo , Homeostase , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Modelos Animais , Mutagênese Sítio-Dirigida , Fosfoenolpiruvato Carboxiquinase (GTP)/metabolismo , Regiões Promotoras Genéticas , Proteínas/genética , Transcriptoma , Dedos de Zinco/genética
18.
Biosci Biotechnol Biochem ; 83(9): 1740-1746, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31021712

RESUMO

Sterol regulatory element-binding proteins (SREBPs) are transcription factors that regulate the expression of genes involved in fatty acid and cholesterol biosynthetic pathways. The present study showed that the flavonoid chrysin impairs the fatty acid synthase promoter. Chrysin reduces the expression of SREBP target genes, such as fatty acid synthase, in human hepatoma Huh-7 cells and impairs de novo synthesis of fatty acids and cholesterol. Moreover, it reduces the endogenous mature, transcriptionally active forms of SREBPs, which are generated by the proteolytic processing of precursor forms. In addition, chrysin reduces the enforced expressing mature forms of SREBPs and their transcriptional activity. The ubiquitin-proteasome system is not involved in the chrysin-mediated reduction of SREBPs mature forms. These results suggest that chrysin suppresses SREBP activity, at least partially, via the degradation of SREBPs mature forms. Abbreviations: ACC1: acetyl-CoA carboxylase 1; DMEM: Dulbecco's modified Eagle's medium; FAS: fatty acid synthase; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; 25-HC: 25-hydroxycholesterol; HMGCS: HMG-CoA synthase; LDH: lactate dehydrogenase; LPDS: lipoprotein-deficient serum; PI3K: phosphatidylinositol 3-kinase; SCD1: stearoyl-CoA desaturase; SREBPs: sterol regulatory element-binding proteins.


Assuntos
Flavonoides/farmacologia , Proteínas de Ligação a Elemento Regulador de Esterol/metabolismo , Animais , Linhagem Celular Tumoral , Colesterol/biossíntese , Ácido Graxo Sintases/genética , Ácidos Graxos/biossíntese , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Regiões Promotoras Genéticas , Proteólise , Proteínas de Ligação a Elemento Regulador de Esterol/antagonistas & inibidores , Proteínas de Ligação a Elemento Regulador de Esterol/genética
19.
Int J Mol Sci ; 20(6)2019 Mar 26.
Artigo em Inglês | MEDLINE | ID: mdl-30917509

RESUMO

An inverse association exists between physical activity and breast cancer incidence and outcomes. An objective indicator of an individual's recent physical activity exposure is aerobic capacity. We took advantage of the fact that there is an inherited as well as inducible component of aerobic capacity to show that experimentally induced mammary cancer is inversely related to inherent aerobic capacity (IAC). The objective of this study was to determine whether cell signaling pathways involved in the development of mammary cancer differed in rats with low inherent aerobic capacity (LIAC, n = 55) versus high inherent aerobic capacity (HIAC, n = 57). Cancer burden was 0.21 ± 0.16 g/rat in HIAC versus 1.14 ± 0.45 in LIAC, p < 0.001. Based on protein expression, cancer in LIAC animals was associated with upregulated glucose utilization, and protein and fatty acid synthesis. Signaling in cancers from HIAC rats was associated with energy sensing, fatty acid oxidation and cell cycle arrest. These findings support the thesis that pro-glycolytic, metabolic inflexibility in LIAC favors not only insulin resistance and obesity but also tumor development and growth. This provides an unappreciated framework for understanding how obesity and low aerobic fitness, hallmarks of physical inactivity, are associated with higher cancer risk and poorer prognosis.


Assuntos
Carcinoma/metabolismo , Neoplasias Mamárias Experimentais/metabolismo , Consumo de Oxigênio , Transdução de Sinais , Animais , Carcinoma/etiologia , Metabolismo Energético , Ácidos Graxos/biossíntese , Feminino , Glucose/metabolismo , Neoplasias Mamárias Experimentais/etiologia , Biossíntese de Proteínas , Ratos
20.
Int J Mol Sci ; 20(6)2019 Mar 18.
Artigo em Inglês | MEDLINE | ID: mdl-30889783

RESUMO

Both cytosolic fatty acid synthesis (FAS) and mitochondrial fatty acid oxidation (FAO) have been shown to play a role in the survival and proliferation of cancer cells. This study aimed to confirm experimentally whether FAS and FAO coexist in breast cancer cells (BCC). By feeding cells with 13C-labeled glutamine and measuring labeling patterns of TCA intermediates, it was possible to show that part of the cytosolic acetyl-CoA used in lipid synthesis is also fed back into the mitochondrion via fatty acid degradation. This results in the transfer of reductive potential from the cytosol (in the form of NADPH) to the mitochondrion (in the form of NADH and FADH2). The hypothesized mechanism was further confirmed by blocking FAS and FAO with siRNAs. Exposure to staurosporine (which induces ROS production) resulted in the disruption of simultaneous FAS and FAO, which could be explained by NADPH depletion.


Assuntos
Ácidos Graxos/biossíntese , Estresse Oxidativo , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Isótopos de Carbono , Linhagem Celular Tumoral , Ácido Graxo Sintases/metabolismo , Feminino , Humanos , Potencial da Membrana Mitocondrial , Análise do Fluxo Metabólico , Oxirredução
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