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1.
Brain Res ; 1827: 148758, 2024 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-38199308

RESUMO

BACKGROUND: Subarachnoid hemorrhage (SAH) is a life-threatening neurological disease that usually has a poor prognosis. Neurogenesis is a potential therapeutic target for brain injury. Ketone metabolism also plays neuroprotective roles in many neurological disorders. OXCT1 (3-Oxoacid CoA-Transferase 1) is the rate-limiting enzyme of ketone body oxidation. In this study, we explored whether increasing ketone oxidation by upregulating OXCT1 in neurons could promote neurogenesis after SAH, and evaluated the potential mechanism involved in this process. METHODS: The ß-hydroxybutyrate content was measured using an enzymatic colorimetric assay. Adeno-associated virus targeting neurons was injected to overexpress OXCT1, and the expression and localization of proteins were evaluated by western blotting and immunofluorescence staining. Adult hippocampal neurogenesis was evaluated by dual staining with doublecortin and 5-Ethynyl-2'-Deoxyuridine. LY294002 was intracerebroventricularly administered to inhibit Akt activity. The Morris water maze and Y-maze tests were employed to assess cognitive function after SAH. RESULTS: The results showed that OXCT1 expression and hippocampal neurogenesis significantly decreased in the early stage of SAH. Overexpression of OXCT1 successfully increased hippocampal neurogenesis via activation of Akt/GSK-3ß/ß-catenin signaling and improved cognitive function, both of which were reversed by administration of LY294002. CONCLUSIONS: OXCT1 regulated hippocampal ketone body metabolism and increased neurogenesis through mechanisms mediated by the Akt/GSK-3ß/ß-catenin pathway, improving cognitive impairment after SAH.


Assuntos
Coenzima A-Transferases , Disfunção Cognitiva , Hipocampo , Neurogênese , Hemorragia Subaracnóidea , Ácido 3-Hidroxibutírico , beta Catenina , Coenzima A-Transferases/genética , Coenzima A-Transferases/metabolismo , Glicogênio Sintase Quinase 3 beta , Hipocampo/crescimento & desenvolvimento , Proteínas Proto-Oncogênicas c-akt , Animais , Camundongos
2.
Sci Rep ; 14(1): 1713, 2024 01 19.
Artigo em Inglês | MEDLINE | ID: mdl-38242911

RESUMO

Ketone bodies serve as an energy source, especially in the absence of carbohydrates or in the extended exercise. Adenosine monophosphate (AMP)-activated protein kinase (AMPK) is a crucial energy sensor that regulates lipid and glucose metabolism. However, whether AMPK regulates ketone metabolism in whole body is unclear even though AMPK regulates ketogenesis in liver. Prolonged resulted in a significant increase in blood and urine levels of ketone bodies in wild-type (WT) mice. Interestingly, fasting AMPKα2-/- and AMPKα1-/- mice exhibited significantly higher levels of ketone bodies in both blood and urine compared to fasting WT mice. BHB tolerance assays revealed that both AMPKα2-/- and AMPKα1-/- mice exhibited slower ketone consumption compared to WT mice, as indicated by higher blood BHB or urine BHB levels in the AMPKα2-/- and AMPKα1-/- mice even after the peak. Interestingly, fasting AMPKα2-/- and AMPKα1-/- mice exhibited significantly higher levels of ketone bodies in both blood and urine compared to fasting WT mice. . Specifically, AMPKα2ΔMusc mice showed approximately a twofold increase in blood BHB levels, and AMPKα2ΔMyo mice exhibited a 1.5-fold increase compared to their WT littermates after a 48-h fasting. However, blood BHB levels in AMPKα1ΔMusc and AMPKα1ΔMyo mice were as same as in WT mice. Notably, AMPKα2ΔMusc mice demonstrated a slower rate of BHB consumption in the BHB tolerance assay, whereas AMPKα1ΔMusc mice did not show such an effect. Declining rates of body weights and blood glucoses were similar among all the mice. Protein levels of SCOT, the rate-limiting enzyme of ketolysis, decreased in skeletal muscle of AMPKα2-/- mice. Moreover, SCOT protein ubiquitination increased in C2C12 cells either transfected with kinase-dead AMPKα2 or subjected to AMPKα2 inhibition. AMPKα2 physiologically binds and stabilizes SCOT, which is dependent on AMPKα2 activity.


Assuntos
Proteínas Quinases Ativadas por AMP , Corpos Cetônicos , Animais , Camundongos , Proteínas Quinases Ativadas por AMP/metabolismo , Jejum , Cetonas , Camundongos Knockout , Ubiquitinação , Coenzima A-Transferases/metabolismo
3.
Molecules ; 29(2)2024 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-38257328

RESUMO

(R)-Benzylsuccinate is generated in anaerobic toluene degradation by the radical addition of toluene to fumarate and further degraded to benzoyl-CoA by a ß-oxidation pathway. Using metabolic modules for benzoate transport and activation to benzoyl-CoA and the enzymes of benzylsuccinate ß-oxidation, we established an artificial pathway for benzylsuccinate production in Escherichia coli, which is based on its degradation pathway running in reverse. Benzoate is supplied to the medium but needs to be converted to benzoyl-CoA by an uptake transporter and a benzoate-CoA ligase or CoA-transferase. In contrast, the second substrate succinate is endogenously produced from glucose under anaerobic conditions, and the constructed pathway includes a succinyl-CoA:benzylsuccinate CoA-transferase that activates it to the CoA-thioester. We present first evidence for the feasibility of this pathway and explore product yields under different growth conditions. Compared to aerobic cultures, the product yield increased more than 1000-fold in anaerobic glucose-fermenting cultures and showed further improvement under fumarate-respiring conditions. An important bottleneck to overcome appears to be product excretion, based on much higher recorded intracellular concentrations of benzylsuccinate, compared to those excreted. While no export system is known for benzylsuccinate, we observed an increased product yield after adding an unspecific mechanosensitive channel to the constructed pathway.


Assuntos
Coenzima A-Transferases , Escherichia coli , Escherichia coli/genética , Succinatos , Benzoatos , Fumaratos , Glucose , Tolueno
4.
Gene ; 896: 148053, 2024 Feb 20.
Artigo em Inglês | MEDLINE | ID: mdl-38042218

RESUMO

Mitochondrial dysfunction plays a critical role in muscular homeostasis, but the molecular mechanism underlying mitochondrial dynamics and sarcopenia awaits to be uncovered. We all know that malnutrition, cachexia, and type 2 diabetes are significant contributors to the development of sarcopenia.Therefore, we analyzed a bioinformatic analysis on cathectic differentially expressed genes (cDEGs), fasted differentially genes (fDEGs) and mitochondria-related genes. The overlapping genes identified were then validated by RT-qPCR and Western blotting experiments in various sarcopenia mice models and used to predict aging-related muscle loss in humans. First, the correlation analysis and PPI network indicated 6 overlapping candidates (Bdh1, Gdap1, Acss1, Mtfp1, Idh2, Oxct1) may constitute a regulatory effect in mitochondrial dynamics and muscle wasting. Next, we successfully established fasted, Lewis lung carcinoma (LLC) and Diabetes Mellitus (DM) induced sarcopenia mice models and verified that Acss1, Mtfp1 and Oxct1 shared common and significant variation tendency in these sarcopenia mice models. Further-more, Pearson correlation analysis showed that Acss1 was negatively related to the weight of gastrocnemius while Mtfp1 and Oxct1 displayed a significantly positive correlation with gastrocnemius weight in sarcopenic mice model induced by LLC, fasting and DM. What's more, ROC analysis based on human aging-related datasets indicated Acss1, Mtfp1, Oxct1 had outstanding diagnostic capabilities for sarcopenia. In general, we identified three hub genes (Acss1, Mtfp1 and Oxct1) that are strongly associated with mitochondrial dysfunction in sarcopenia and may provide novel and reliable indicators for screening, diagnosis, and prognosis, as well as potential therapeutic targets for patients with sarcopenia.


Assuntos
Diabetes Mellitus Tipo 2 , Doenças Mitocondriais , Sarcopenia , Animais , Humanos , Camundongos , Envelhecimento/genética , Biomarcadores , Diabetes Mellitus Tipo 2/genética , Sarcopenia/diagnóstico , Sarcopenia/genética , Sarcopenia/patologia , Coenzima A-Transferases
5.
Sci Rep ; 13(1): 14413, 2023 09 02.
Artigo em Inglês | MEDLINE | ID: mdl-37660163

RESUMO

Long-term effect of Direct-acting antivirals (DAAs) on gut microbiota, short-chain fatty acids (SCFAs) and microbial translocation in patients with hepatitis C virus (HCV) infection who achieve sustained virological response (SVR) were limited. A longitudinal study of 50 patients with HCV monoinfection and 19 patients with HCV/HIV coinfection received DAAs were conducted. Fecal specimens collected at baseline and at week 72 after treatment completion (FUw72) were analyzed for 16S rRNA sequencing and the butyryl-CoA:acetateCoA transferase (BCoAT) gene expression using real-time PCR. Plasma lipopolysaccharide binding protein (LBP) and intestinal fatty acid binding protein (I-FABP) were quantified by ELISA assays. SVR rates in mono- and coinfected patients were comparable (94% vs. 100%). The improvement of gut dysbiosis and microbial translocation was found in responders but was not in non-responders. Among responders, significant restoration of alpha-diversity, BCoAT and LBP were observed in HCV patients with low-grade fibrosis (F0-F1), while HCV/HIV patients exhibited partial improvement at FUw72. I-FABP did not decline significantly in responders. Treatment induced microbiota changes with increasing abundance of SCFAs-producing bacteria, including Blautia, Fusicatenibacter, Subdoligranulum and Bifidobacterium. In conclusion, long-term effect of DAAs impacted the restoration of gut dysbiosis and microbial translocation. However, early initiation of DAAs required for an alteration of gut microbiota, enhanced SCFAs-producing bacteria, and could reduce HCV-related complications.


Assuntos
Infecções por HIV , Hepatite C Crônica , Hepatite C , Humanos , Hepacivirus/genética , Antivirais/uso terapêutico , Disbiose/complicações , Infecções por HIV/complicações , Infecções por HIV/tratamento farmacológico , Estudos Longitudinais , RNA Ribossômico 16S , Hepatite C Crônica/complicações , Hepatite C Crônica/tratamento farmacológico , Hepatite C/complicações , Hepatite C/tratamento farmacológico , Clostridiales , Coenzima A-Transferases
6.
Rev Med Inst Mex Seguro Soc ; 61(5): 691-694, 2023 Sep 04.
Artigo em Espanhol | MEDLINE | ID: mdl-37773183

RESUMO

Background: Succinyl-CoA:3 oxoacid CoA transferase deficiency (SCOTD) is a rare autosomal recessive disease, characterized by altered utilization of ketone bodies, with acute episodes of ketoacidosis. Clinical case: It is presented the case of a patient with SCOTD, with a first atypical episode accompanied by hyperglycemia, with 4 subsequent episodes with classic manifestations of the disease, presenting with a biochemical pattern of permanent ketonuria with marked elevation of ketone bodies (acetoacetate, 3 beta-hydroxybutyrate) in the study of urinary organic acids by gas chromatography and mass spectrometry, together with the clinical picture granting the diagnosis. It was started a maintenance therapy with a characteristic feeding plan; it was shown an adequate response to treatment, and the absence of permanent ketosis was surmised. Conclusion: Being a rare disease, the categorization of these patients as diabetic ketoacidosis is frequent. The clinical and biochemical characteristics with ketosis or persistent ketonuria should be analyzed very carefully, especially in patients presenting with hyperglycemia, which is an atypical manifestation of the disease, in order to make an early diagnosis and treatment, positively impacting the prognosis of patients.


Introducción: la deficiencia de succinil-CoA acetoacetato transferasa (SCOT) es una enfermedad rara, autosómica recesiva, caracterizada por alteración en la utilización de cuerpos cetónicos, con episodios agudos de cetoacidosis. Caso clínico: se presenta el caso de un paciente con deficiencia de SCOT, con un primer episodio atípico acompañado con hiperglucemia, con 4 episodios posteriores con manifestaciones clásicas de la enfermedad, que presentó patrón bioquímico de cetonuria permanente con marcada elevación de cuerpos cetónicos (acetoacetato, 3 beta-hidroxibutirato) en estudio de ácidos orgánicos urinarios por cromatografía de gases y espectrometría de masas, aunado a cuadro clínico que otorgó el diagnóstico. Se inició terapia de mantenimiento con plan de alimentación característico; se demostró una adecuada respuesta al tratamiento, y se infirió una ausencia de cetosis permanente. Conclusiones: al ser una enfermedad rara, la categorización de estos pacientes como cetoacidosis diabética es frecuente. Se deben analizar de forma muy minuciosa las características clínicas y bioquímicas con cetosis o cetonuria persistente, sobre todo en pacientes que se presenten con hiperglucemia, que es una manifestación atípica de la enfermedad, para realizar un diagnóstico y tratamiento temprano que impacte de forma positiva el pronóstico de los pacientes.


Assuntos
Hiperglicemia , Cetose , Humanos , Coenzima A-Transferases , Corpos Cetônicos , Cetose/etiologia , Ácido 3-Hidroxibutírico/análise , Hiperglicemia/complicações
8.
Methods Enzymol ; 683: 19-39, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37087187

RESUMO

BAHD acyl-coenzyme A (CoA) acyltransferases play key roles in a large number of biosynthetic reactions involved in plant specialized metabolism. One approach to measure reaction rates for these enzymes is to quantify the amide or ester reaction products following chromatographic separation of reaction components, an approach that can be labor intensive and time consuming, and complicated by a lack of pure standards. We previously developed and validated an alternative approach using 5,5'-dithio-bis-(2-nitrobenzoic acid) (DTNB, Ellman's reagent) to spectrophotometrically monitor reaction progress by the release of free CoA in the reaction. This approach allows near-real time measurement of reaction rates, permitting reaction conditions (buffer, reactant, and enzyme concentrations, etc.) to be changed "on the fly." The ease and rapidity of data collection allows a high density of data points to be collected for determination of kinetic parameters. Here we provide a detailed procedure for using DTNB to measure BAHD acyl-CoA acyltransferase reaction rates, and as an example, use it to determine kinetic parameters for red clover hydroxycinnamoyl-CoA:shikimate hydroxycinnamoyltransferase, a BAHD acyl-CoA hydroxycinnamoyltransferase not previously characterized with respect to kinetic parameters. This approach may be more generally applicable to transferases using CoA donors.


Assuntos
Aciltransferases , Coenzima A-Transferases , Ácido Ditionitrobenzoico/química , Aciltransferases/metabolismo
10.
Diabetes ; 72(1): 126-134, 2023 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-36256885

RESUMO

Despite significant progress in understanding the pathogenesis of type 2 diabetes (T2D), the condition remains difficult to manage. Hence, new therapeutic options targeting unique mechanisms of action are required. We have previously observed that elevated skeletal muscle succinyl CoA:3-ketoacid CoA transferase (SCOT) activity, the rate-limiting enzyme of ketone oxidation, contributes to the hyperglycemia characterizing obesity and T2D. Moreover, we identified that the typical antipsychotic agent pimozide is a SCOT inhibitor that can alleviate obesity-induced hyperglycemia. We now extend those observations here, using computer-assisted in silico modeling and in vivo pharmacology studies that highlight SCOT as a noncanonical target shared among the diphenylbutylpiperidine (DPBP) drug class, which includes penfluridol and fluspirilene. All three DPBPs tested (pimozide, penfluridol, and fluspirilene) improved glycemia in obese mice. While the canonical target of the DPBPs is the dopamine 2 receptor, studies in obese mice demonstrated that acute or chronic treatment with a structurally unrelated antipsychotic dopamine 2 receptor antagonist, lurasidone, was devoid of glucose-lowering actions. We further observed that the DPBPs improved glycemia in a SCOT-dependent manner in skeletal muscle, suggesting that this older class of antipsychotic agents may have utility in being repurposed for the treatment of T2D.


Assuntos
Antipsicóticos , Diabetes Mellitus Tipo 2 , Hiperglicemia , Animais , Camundongos , Antipsicóticos/farmacologia , Antipsicóticos/uso terapêutico , Coenzima A-Transferases , Diabetes Mellitus Tipo 2/tratamento farmacológico , Dopamina , Fluspirileno/farmacologia , Hiperglicemia/tratamento farmacológico , Camundongos Obesos , Penfluridol/farmacologia , Pimozida/farmacologia , Receptores Dopaminérgicos/metabolismo
11.
Biochemistry ; 62(1): 75-84, 2023 01 03.
Artigo em Inglês | MEDLINE | ID: mdl-36535006

RESUMO

Mesaconyl-CoA transferase (Mct) is one of the key enzymes of the 3-hydroxypropionate (3HP) bi-cycle for autotrophic CO2 fixation. Mct is a family III/Frc family CoA transferase that catalyzes an unprecedented intra-molecular CoA transfer from the C1-carboxyl group to the C4-carboxyl group of mesaconate at catalytic efficiencies >106 M-1 s-1. Here, we show that the reaction of Mct proceeds without any significant release of free CoA or the transfer to external acceptor acids. Mct catalyzes intra-molecular CoA transfers at catalytic efficiencies that are at least more than 6 orders of magnitude higher compared to inter-molecular CoA transfers, demonstrating that the enzyme exhibits exquisite control over its reaction. To understand the molecular basis of the intra-molecular CoA transfer in Mct, we solved crystal structures of the enzyme from Chloroflexus aurantiacus in its apo form, as well as in complex with mesaconyl-CoA and several covalently enzyme-bound intermediates of CoA and mesaconate at the catalytically active residue Asp165. Based on these structures, we propose a reaction mechanism for Mct that is similar to inter-molecular family III/Frc family CoA transferases. However, in contrast to the latter that undergo opening and closing cycles during the reaction to exchange substrates, the central cavity of Mct remains sealed ("corked-up") by the CoA moiety, strongly favoring the intra-molecular CoA transfer between the C1 and the C4 position of mesaconate.


Assuntos
Acil Coenzima A , Coenzima A-Transferases
12.
J Biotechnol ; 359: 29-34, 2022 Nov 20.
Artigo em Inglês | MEDLINE | ID: mdl-36150604

RESUMO

Isopropanol has a good potential as a new fuel substitution. In the model biosynthesis pathway of isopropanol synthesis, acetoacetyl-CoA is converted to acetoacetate by acetoacetyl-CoA transferases, which requires an acetate molecule as a substrate. Herein, a novel isopropanol synthesis pathway based on mammalian ketone metabolic pathway was developed. In this pathway, acetoacetyl-CoA is condensed with acetyl-CoA to generate 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) by HMG-CoA synthase, and then catalyzed by HMG-CoA lyase to generate acetoacetate. This process is acetate-independent. Under the same experimental system using glycerol as carbon source, the E. coli strain MG::ISOP1 containing the novel pathway produced 11.7 times more isopropanol than the strain MG::ISOP0 containing the model pathway. The pta-ackA knockout mutant strain MG∆pta-ackA::ISOP1, which reduced the conversion of acetyl-CoA to acetate, further increased the production from 76 mg/L to 360 mg/L. In another strategy, knocking out atoDA to block the acetoacetate degradation pathway in strain MG∆atoDA::ISOP1 increased the production to 680 mg/L. By knocking out both of pta-ackA and atoDA, strain MGΔpta-ackAΔatoDA::ISOP1 produced 964 mg/L of isopropanol, which was 12.7 times that of MG::ISOP1. This study indicated that the novel pathway is competent for isopropanol synthesis, and provides a new perspective for biosynthesis of isopropanol.


Assuntos
2-Propanol , Escherichia coli , Escherichia coli/genética , Escherichia coli/metabolismo , 2-Propanol/metabolismo , Acetoacetatos/metabolismo , Acetilcoenzima A/metabolismo , Coenzima A-Transferases/metabolismo , Hidroximetilglutaril-CoA Sintase/genética , Hidroximetilglutaril-CoA Sintase/metabolismo , Glicerol/metabolismo , Acetatos/metabolismo , Carbono/metabolismo
13.
Appl Microbiol Biotechnol ; 106(19-20): 6861-6876, 2022 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-36071291

RESUMO

This study aimed to characterize the prokaryotic community and putative microbial interactions involved in hydrogen (H2) production during the dark fermentation (DF) process, applying principal components analysis (PCA) to correlate changes in operational, physicochemical, and biological variables. For this purpose, a continuous stirred-tank reactor-type digester fed with tequila vinasses was operated at 24, 18, and 12 h of hydraulic retention times (HRTs) to apply organic loading rates of 20, 36, and 54 g-COD L-1 d-1, corresponding to stages I, II, and III, respectively. Results indicated high population dynamics for Archaea during the DF process toward a decrease in total sequences from 6299 to 99. Concerning the Bacteria community, lactic acid bacteria (LAB) were dominant reaching a relative abundance of 57.67%, while dominant H2-producing bacteria (HPB) decreased from 25.76% to 21.06% during stage III. Putative competitive exclusion mechanisms such as competition for substrates, bacteriocins production, and micronutrient depletion carried out by Archaea and non-H2-producing bacteria (non-HPB), especially LAB, could negatively impact the dominance of HPB such as Ethanoligenens harbinense and Clostridium tyrobutyricum. As a consequence, low maximal volumetric H2 production rate (672 mL-H2 L-1 d-1) and yield (3.88 mol-H2 assimilated sugars-1) were obtained. The global scenario obtained by PCA correlations suggested that C. tyrobutyricum positively impacted H2 molar yield through butyrate fermentation using the butyryl-CoA:acetate CoA transferase pathway, while the most abundant HPB E. harbinense decreased its relative abundance at the shortest HRT toward the dominance of non-HPB. This study provides new insights into the microbial interactions and helps to better understand the DF performance for H2 production using tequila vinasses as substrate. KEY POINTS: • E. harbinense and C. tyrobutyricum were responsible for H2 production. • Clostridiales used acetate and butyrate fermentations for H2 production. • LAB won the competition for sugars against Clostridiales during DF. • Putative bacteriocins production and micronutrients depletion could favor LAB.


Assuntos
Bacteriocinas , Reatores Biológicos , Acetatos/metabolismo , Archaea/metabolismo , Bactérias/genética , Bactérias/metabolismo , Bacteriocinas/metabolismo , Reatores Biológicos/microbiologia , Butiratos/metabolismo , Coenzima A-Transferases/metabolismo , Fermentação , Hidrogênio/metabolismo , Interações Microbianas , Micronutrientes/metabolismo , Açúcares/metabolismo
14.
Diabetes Obes Metab ; 24(11): 2263-2272, 2022 11.
Artigo em Inglês | MEDLINE | ID: mdl-35801343

RESUMO

AIM: To investigate cardiac signalling pathways connecting substrate utilization with left ventricular remodelling in a murine pressure overload model. METHODS: Cardiac hypertrophy was induced by transverse aortic constriction surgery in 20-week-old C57BL/6J mice treated with or without the sodium-glucose co-transporter 2 (SGLT2) inhibitor ertugliflozin (225 mg kg-1 chow diet) for 10 weeks. RESULTS: Ertugliflozin improved left ventricular function and reduced myocardial fibrosis. This occurred simultaneously with a fasting-like response characterized by improved glucose tolerance and increased ketone body concentrations. While cardiac insulin signalling was reduced in response to SGLT2 inhibition, AMP-activated protein kinase (AMPK) signalling was increased with induction of the fatty acid transporter cluster of differentiation 36 and phosphorylation of acetyl-CoA carboxylase (ACC). Further, enzymes responsible for ketone body catabolism (ß-hydroxybutyrate dehydrogenase, succinyl-CoA:3-oxoacid-CoA transferase and acetyl-CoA acetyltransferase 1) were induced by SGLT2 inhibition. Ertugliflozin led to more cardiac abundance of fatty acids, tricarboxylic acid cycle metabolites and ATP. Downstream mechanistic target of rapamycin (mTOR) pathway, relevant for protein synthesis, cardiac hypertrophy and adverse cardiac remodelling, was reduced by SGLT2 inhibition, with alleviation of endoplasmic reticulum (ER) stress and unfolded protein response (UPR) providing a potential mechanism for abundant reduced left ventricular apoptosis and fibrosis. CONCLUSION: SGLT2 inhibition reduced left ventricular fibrosis in a murine model of cardiac hypertrophy. Mechanistically, this was associated with reduced cardiac insulin and increased AMPK signalling as a potential mechanism for less cardiac mTOR activation with alleviation of downstream ER stress, UPR and apoptosis.


Assuntos
Insulinas , Inibidores do Transportador 2 de Sódio-Glicose , Proteínas Quinases Ativadas por AMP/metabolismo , Acetil-CoA C-Acetiltransferase/metabolismo , Acetil-CoA Carboxilase/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Apoptose , Compostos Bicíclicos Heterocíclicos com Pontes , Cardiomegalia/metabolismo , Cardiomegalia/patologia , Coenzima A-Transferases/metabolismo , Estresse do Retículo Endoplasmático , Ácidos Graxos/metabolismo , Fibrose , Glucose/metabolismo , Hidroxibutirato Desidrogenase/metabolismo , Cetoácidos/metabolismo , Cetonas/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Miócitos Cardíacos/metabolismo , Sirolimo/metabolismo , Sódio/metabolismo , Transportador 2 de Glucose-Sódio/metabolismo , Inibidores do Transportador 2 de Sódio-Glicose/farmacologia , Inibidores do Transportador 2 de Sódio-Glicose/uso terapêutico , Serina-Treonina Quinases TOR/metabolismo
15.
Lett Appl Microbiol ; 75(4): 844-856, 2022 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-35575477

RESUMO

Butyrate-producing bacteria generate butyrate, which has antidepressant effects. Xiaoyaosan (XYS), a traditional Chinese medicine (TCM) used to treat depression, may improve depression-like behaviour by modulating the gut microbiota. However, the functional groups and mechanisms of action in the XYS treatment of depression remain unknown. This study aimed to analyse with clone sequencing the changes in intestinal butyrate-producing bacteria in XYS-treated chronic unpredictable mild stress (CUMS) rats. We successfully established the XYS-treated CUMS rat model of depression. Rat faecal samples were collected before, during, and after the experiment, and butyryl-CoA:acetate CoA-transferase gene primers were selected for PCR amplification to determine the diversity of butyrate-producing bacteria. The results showed that XYS increased intestinal butyrate-producing bacterial diversity in CUMS rats regarding phylum and genus numbers; the number of phyla increased to two, distributed in Firmicutes and Bacteroides, and four genera were distributed in Eubacterium sp., Roseburia sp., Clostridium sp. and Bacteroides sp. Only one phylum and two genera were present in the model group without XYS treatment. Our findings indicate that XYS can improve depression-like behaviour by regulating intestinal butyrate-producing bacteria diversity, particularly Roseburia sp. and Eubacterium sp., thus providing new insights into the targeted regulation of the intestinal flora to treat depression.


Assuntos
Coenzima A-Transferases , Depressão , Acetatos , Animais , Antidepressivos/farmacologia , Bactérias , Comportamento Animal , Butiratos/farmacologia , Coenzima A-Transferases/farmacologia , Depressão/tratamento farmacológico , Depressão/genética , Depressão/microbiologia , Modelos Animais de Doenças , Medicamentos de Ervas Chinesas , Ratos
16.
Cell ; 185(3): 513-529.e21, 2022 02 03.
Artigo em Inglês | MEDLINE | ID: mdl-35120663

RESUMO

The human gut microbiota resides within a diverse chemical environment challenging our ability to understand the forces shaping this ecosystem. Here, we reveal that fitness of the Bacteroidales, the dominant order of bacteria in the human gut, is an emergent property of glycans and one specific metabolite, butyrate. Distinct sugars serve as strain-variable fitness switches activating context-dependent inhibitory functions of butyrate. Differential fitness effects of butyrate within the Bacteroides are mediated by species-level variation in Acyl-CoA thioesterase activity and nucleotide polymorphisms regulating an Acyl-CoA transferase. Using in vivo multi-omic profiles, we demonstrate Bacteroides fitness in the human gut is associated together, but not independently, with Acyl-CoA transferase expression and butyrate. Our data reveal that each strain of the Bacteroides exists within a unique fitness landscape based on the interaction of chemical components unpredictable by the effect of each part alone mediated by flexibility in the core genome.


Assuntos
Microbioma Gastrointestinal , Metaboloma , Polissacarídeos/metabolismo , Acil Coenzima A/metabolismo , Sequência de Aminoácidos , Aminoácidos de Cadeia Ramificada/metabolismo , Bacteroidetes/efeitos dos fármacos , Bacteroidetes/genética , Bacteroidetes/crescimento & desenvolvimento , Butiratos/química , Butiratos/farmacologia , Coenzima A-Transferases/química , Coenzima A-Transferases/metabolismo , Microbioma Gastrointestinal/efeitos dos fármacos , Microbioma Gastrointestinal/genética , Variação Genética/efeitos dos fármacos , Concentração de Íons de Hidrogênio , Metaboloma/efeitos dos fármacos , Metaboloma/genética , Polimorfismo de Nucleotídeo Único/genética , Regiões Promotoras Genéticas/genética , Especificidade da Espécie , Estresse Fisiológico/efeitos dos fármacos , Estresse Fisiológico/genética , Transcrição Gênica/efeitos dos fármacos
17.
Protein Sci ; 31(4): 864-881, 2022 04.
Artigo em Inglês | MEDLINE | ID: mdl-35049101

RESUMO

The coenzyme A (CoA) transferases are a superfamily of proteins central to the metabolism of acetyl-CoA and other CoA thioesters. They are diverse group, catalyzing over a 100 biochemical reactions and spanning all three domains of life. A deeply rooted idea, proposed two decades ago, is these enzymes fall into three families (I, II, and III). Here we find they fall into different families, which we achieve by analyzing all CoA transferases characterized to date. We manually annotated 94 CoA transferases with functional information (including rates of catalysis for 208 reactions) from 97 publications. This represents all enzymes we could find in the primary literature, and it is double the number annotated in four protein databases (BRENDA, KEGG, MetaCyc, UniProt). We found family I transferases are not closely related to each other in terms of sequence, structure, and reactions catalyzed. This family is not even monophyletic. These problems are solved by regrouping the three families into six, including one family with many non-CoA transferases. The problem (and solution) became apparent only by analyzing our large set of manually annotated proteins. It would have been missed if we had used the small number of proteins annotated in UniProt and other databases. Our work is important to understanding the biology of CoA transferases. It also warns investigators doing phylogenetic analyses of proteins to go beyond information in databases.


Assuntos
Proteínas de Bactérias , Coenzima A-Transferases , Proteínas de Bactérias/química , Catálise , Coenzima A , Coenzima A-Transferases/química , Coenzima A-Transferases/genética , Coenzima A-Transferases/metabolismo , Bases de Dados de Proteínas , Humanos , Filogenia
18.
Microb Cell Fact ; 20(1): 229, 2021 Dec 23.
Artigo em Inglês | MEDLINE | ID: mdl-34949197

RESUMO

BACKGROUND: Steroid drugs are essential for disease prevention and clinical treatment. However, due to intricated steroid structure, traditional chemical methods are rarely implemented into the whole synthetic process for generating steroid intermediates. Novel steroid drug precursors and their ideal bacterial strains for industrial production have yet to be developed. Among these, 9,21-dihydroxy-20-methyl-pregna-4-en-3-one (9-OH-4-HP) is a novel steroid drug precursor, suitable for the synthesis of corticosteroids. In this study, a combined strategy of blocking Δ1-dehydrogenation and the C19 pathway as well as improving the intracellular environment was investigated to construct an effective 9-OH-4-HP-producing strain. RESULTS: The Δ1-dehydrogenation-deficient strain of wild-type Mycobacterium neoaurum DSM 44074 produces 9-OH-4-HP with a molar yield of 4.8%. Hsd4A, encoding a ß-hydroxyacyl-CoA dehydrogenase, and fadA5, encoding an acyl-CoA thiolase, were separately knocked out to block the C19 pathway in the Δ1-dehydrogenation-deficient strain. The two engineered strains were able to accumulate 0.59 g L-1 and 0.47 g L-1 9-OH-4-HP from 1 g L-1 phytosterols, respectively. Furthermore, hsd4A and fadA5 were knocked out simultaneously in the Δ1-dehydrogenation-deficient strain. The 9-OH-4-HP production from the Hsd4A and FadA5 deficient strain was 11.9% higher than that of the Hsd4A deficient strain and 40.4% higher than that of the strain with FadA5 deficiency strain, respectively. The purity of 9-OH-4-HP obtained from the Hsd4A and FadA5 deficient strain has reached 94.9%. Subsequently, the catalase katE from Mycobacterium neoaurum and an NADH oxidase, nox, from Bacillus subtilis were overexpressed to improve the intracellular environment, leading to a higher 9-OH-4-HP production. Ultimately, 9-OH-4-HP production reached 3.58 g L-1 from 5 g L-1 phytosterols, and the purity of 9-OH-4-HP improved to 97%. The final 9-OH-4-HP production strain showed the best molar yield of 85.5%, compared with the previous reported strain with 30% molar yield of 9-OH-4-HP. CONCLUSION: KstD, Hsd4A, and FadA5 are key enzymes for phytosterol side-chain degradation in the C19 pathway. Double deletion of hsd4A and fadA5 contributes to the blockage of the C19 pathway. Improving the intracellular environment of Mycobacterium neoaurum during phytosterol bioconversion could accelerate the conversion process and enhance the productivity of target sterol derivatives.


Assuntos
Redes e Vias Metabólicas , Mycobacteriaceae/genética , Mycobacteriaceae/metabolismo , Fitosteróis/metabolismo , Pró-Fármacos/metabolismo , Esteroides/metabolismo , Proteínas de Bactérias/genética , Coenzima A-Transferases/genética , Edição de Genes , Técnicas de Inativação de Genes , Genoma Bacteriano , Hidroliases/genética , Oxirredutases/genética
19.
PLoS One ; 16(11): e0260116, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34797858

RESUMO

Lactobacillus paragasseri was identified as a novel sister taxon of L. gasseri in 2018. Since the reclassification of L. paragasseri, there has been hardly any report describing the probiotic properties of this species. In this study, an L. paragasseri strain UBLG-36 was sequenced and analyzed to determine the molecular basis that may confer the bacteria with probiotic potential. UBLG-36 was previously documented as an L. gasseri strain. Average nucleotide identity and phylogenomic analysis allowed accurate taxonomic identification of UBLG-36 as an L. paragasseri strain. Analysis of the draft genome (~1.94 Mb) showed that UBLG-36 contains 5 contigs with an average G+C content of 34.85%. Genes essential for the biosynthesis of bacteriocins, adhesion to host epithelium, stress resistance, host immunomodulation, defense, and carbohydrate metabolism were identified in the genome. Interestingly, L. paragasseri UBLG-36 also harbored genes that code for enzymes involved in oxalate catabolism, such as formyl coenzyme A transferase (frc) and oxalyl coenzyme A decarboxylase (oxc). In vitro oxalate degradation assay showed that UBLG-36 is highly effective in degrading oxalate (averaging more than 45% degradation), a feature that has not been reported before. As a recently identified bacterium, there are limited genomic reports on L. paragasseri, and our draft genome sequence analysis is the first to describe and emphasize the probiotic potential and oxalate degrading ability of this species. With results supporting the probiotic functionalities and oxalate catabolism of UBLG-36, we propose that this strain is likely to have immense biotechnological applications upon appropriate characterization.


Assuntos
Lactobacillus/genética , Lactobacillus/metabolismo , Oxalatos/metabolismo , Bacteriocinas/metabolismo , Composição de Bases/genética , Carboxiliases/metabolismo , Coenzima A-Transferases/metabolismo , Genômica , Imunomodulação , Nucleotídeos , Filogenia , Probióticos/metabolismo , Análise de Sequência , Sequenciamento Completo do Genoma/métodos
20.
Am J Physiol Heart Circ Physiol ; 321(4): H751-H755, 2021 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-34533402

RESUMO

Lack of glucose uptake compromises metabolic flexibility and reduces energy efficiency in the diabetes mellitus (DM) heart. Although increased use of fatty acid to compensate glucose substrate has been studied, less is known about ketone body metabolism in the DM heart. Ketogenic diet reduces obesity, a risk factor for T2DM. How ketogenic diet affects ketone metabolism in the DM heart remains unclear. At the metabolic level, the DM heart differs from the non-DM heart because of altered metabolic substrate and the T1DM heart differs from the T2DM heart because of insulin levels. How these changes affect ketone body metabolism in the DM heart are poorly understood. Ketogenesis produces ketone bodies by using acetyl-CoA, whereas ketolysis consumes ketone bodies to produce acetyl-CoA, showing their opposite roles in the ketone body metabolism. Cardiac-specific transgenic upregulation of ketogenesis enzyme or knockout of ketolysis enzyme causes metabolic abnormalities leading to cardiac dysfunction. Empirical evidence demonstrates upregulated transcription of ketogenesis enzymes, no change in the levels of ketone body transporters, very high levels of ketone bodies, and reduced expression and activity of ketolysis enzymes in the T1DM heart. Based on these observations, I hypothesize that increased transcription and activity of cardiac ketogenesis enzyme suppresses ketolysis enzyme in the DM heart, which decreases cardiac energy efficiency. The T1DM heart exhibits highly upregulated ketogenesis compared with the T2DM heart because of the lack of insulin, which inhibits ketogenesis enzyme.


Assuntos
Glicemia/metabolismo , Diabetes Mellitus Tipo 1/complicações , Diabetes Mellitus Tipo 2/complicações , Cardiomiopatias Diabéticas/etiologia , Metabolismo Energético , Insulina/metabolismo , Corpos Cetônicos/metabolismo , Miocárdio/metabolismo , Animais , Coenzima A-Transferases/genética , Coenzima A-Transferases/metabolismo , Diabetes Mellitus Tipo 1/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Cardiomiopatias Diabéticas/metabolismo , Cetoacidose Diabética/etiologia , Cetoacidose Diabética/metabolismo , Dieta Cetogênica , Feminino , Humanos , Hidroximetilglutaril-CoA Sintase/genética , Hidroximetilglutaril-CoA Sintase/metabolismo , Masculino
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