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1.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi ; 38(2): 166-169, 2021 Feb 10.
Artigo em Chinês | MEDLINE | ID: mdl-33565073

RESUMO

OBJECTIVE: To explore the genetic etiology of a child suspected for ß-ketothiolase deficiency by neonatal screening. METHODS: All coding exons and flanking sequences of the ACAT1 gene were subjected to targeted capture and high-throughput sequencing. Suspected variants were verified by Sanger sequencing and bioinformatic analysis. RESULTS: The child was found to harbor compound heterozygous variants of the ACAT1 gene, namely c.121-3C>G and c.275G>A (p. Gly92Asp). The c.121-3C>G variant was also detected in his father and two sisters, while the c.275G>A (p. Gly92Asp) was a de novo variant. A c.334+ 172C>G (rs12226047) polymorphism was also detected in his mother and two sisters. Sanger sequencing has verified that the c.275G>A (p. Gly92Asp) and c.334+172C>G (rs12226047) variants are located on the same chromosome. Bioinformatics analysis suggested both c.121-3C>G and c.275G>A (p.G92D) variants to be damaging. Based on the American College of Medical Genetics and Genomics standards and guidelines, the c.275G>A variant of the ACAT1 gene was predicted to be pathogenic (PS2+ PM2+ PM3+ PP3+PP4), the c.121-3C>G variant to be likely pathogenic (PM2+ PM3+ PP3+PP4). CONCLUSION: The c.121-3C>G and c.275G>A variants of the ACAT1 gene probably underlay the pathogenesis of the child. Above finding has enriched the variant spectrum of the ACAT1 gene.


Assuntos
Acetil-CoA C-Acetiltransferase/genética , Acetil-CoA C-Aciltransferase/deficiência , Erros Inatos do Metabolismo dos Aminoácidos/genética , Acetil-CoA C-Aciltransferase/genética , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Recém-Nascido , Masculino , Mutação
2.
J Biosci Bioeng ; 130(3): 290-294, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32482608

RESUMO

Aeromonas hydrophila 4AK4 normally produces the copolymer of 3-hydroxybutyrate and 3-hydroxyhexanoate (PHBHHx) using lauric acid as the carbon source. In this study we reported the metabolic engineering of A. hydrophila 4AK4 for the production of polyhydroxyalkanoate (PHA) using acetate as a main carbon source. Recombinant A. hydrophila overexpressing ß-ketothiolase and acetoacetyl-CoA reductase could accumulate poly-3-hydroxybutyrate (PHB) from acetate with a polymer content of 1.39 wt%. Further overexpression of acetate kinase/phosphotransacetylase and acetyl-CoA synthetase improved PHB content to 8.75 wt% and 19.82 wt%, respectively. When acetate and propionate were simultaneously supplied as carbon sources, the engineered A. hydrophila overexpressing ß-ketothiolase, acetoacetyl-CoA reductase, and acetyl-CoA synthetase was found able to produce the copolymer of 3-hydroxybutyrate and 3-hydroxyvalerate (PHBV). The recombinant grew to 3.79 g/L cell dry weight (CDW) containing 15.02 wt% PHBV. Our proposed metabolic engineering strategies illustrate the feasibility for producing PHA from acetate by A. hydrophila.


Assuntos
Acetatos/metabolismo , Aeromonas hydrophila/genética , Aeromonas hydrophila/metabolismo , Engenharia Metabólica , Poli-Hidroxialcanoatos/biossíntese , Ácido 3-Hidroxibutírico/metabolismo , Acetil-CoA C-Aciltransferase/genética , Oxirredutases do Álcool/genética , Ácidos Pentanoicos/metabolismo
3.
Biochim Biophys Acta Gene Regul Mech ; 1862(10): 194436, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31682939

RESUMO

Histone H2A.Z plays an essential role in regulating transcriptional rates and memory. Interestingly, H2A.Z-bound nucleosomes are located in both transcriptionally active and inactive promotors, with no clear understanding of the mechanisms via which it differentially regulates transcription. We hypothesized that its functions are mediated through recruitment of regulatory proteins to promoters. Using rapid chromatin immunoprecipitation-mass spectrometry, we uncovered the association of H2A.Z-bound chromatin with the metabolic enzymes, oxoglutarate dehydrogenase (OGDH) and acetyl-CoA acyltransferase 2 (ACAA2). Recombinant green florescence fusion proteins, combined with mutations of predicted nuclear localization signals, confirmed their nuclear localization and chromatin binding. Conclusively, chromatin immunoprecipitation-deep sequencing, confirmed the predominant association of OGDH and ACAA2 with H2A.Z-occupied transcription start sites and enhancers, the former of which we confirmed is conserved in both mouse and human tissue. Furthermore, H2A.Z-deficient human HAP1 cells exhibited reduced chromatin-bound metabolic enzymes, accompanied with reduced posttranslational histone modifications, including acetylation and succinylation. Specifically, knockdown of OGDH diminished H4 succinylation. Thus, the data reveal that select metabolic enzymes are assembled at active, H2A.Z-occupied, promoters, for potential site-directed production of metabolic intermediates that are required for histone modifications.


Assuntos
Acetilcoenzima A/genética , Acetil-CoA C-Aciltransferase/genética , Histonas/genética , Complexo Cetoglutarato Desidrogenase/genética , Acetilação , Animais , Cromatina/genética , Código das Histonas/genética , Humanos , Camundongos , Proteínas do Tecido Nervoso/genética , Nucleossomos/genética , Regiões Promotoras Genéticas , Processamento de Proteína Pós-Traducional/genética , Fatores de Transcrição/genética , Sítio de Iniciação de Transcrição
4.
Free Radic Biol Med ; 145: 349-356, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31605749

RESUMO

Appropriate diet is essential for the regulation of age-related macular degeneration (AMD). In particular the type of dietary polyunsaturated fatty acids (PUFA) and poor antioxidant status including carotenoid levels concomitantly contribute to AMD risk. Build-up of oxidative stress in AMD induces PUFA oxidation, and a mix of lipid oxidation products (LOPs) are generated. However, LOPs are not comprehensively evaluated in AMD. LOPs are considered biomarkers of oxidative stress but also contributes to inflammatory response. In this cross-sectional case-control study, plasma omega-6/omega-3 PUFA ratios and antioxidant status (glutathione, superoxide dismutase and catalase), and plasma and urinary LOPs (41 types) were determined to evaluate its odds-ratio in the risk of developing exudative AMD (n = 99) compared to age-gender-matched healthy controls (n = 198) in adults with Chinese diet. The odds ratio of developing exudative AMD increased with LOPs from omega-6 PUFA and decreased from those of omega-3 PUFA. These observations were associated with a high plasma omega-6/omega-3 PUFA ratio and low carotenoid levels. In short, poor PUFA and antioxidant status increased the production of omega-6 PUFA LOPs such as dihomo-isoprostane and dihomo-isofuran, and lowered omega-3 PUFA LOPs such as neuroprostanes due to the high omega-6/omega-3 PUFA ratios; they were also correlated to the risk of AMD development. These findings indicate the generation of specific LOPs is associated with the development of exudative AMD.


Assuntos
Ácidos Graxos Ômega-3/administração & dosagem , Ácidos Graxos Ômega-6/administração & dosagem , Degeneração Macular/metabolismo , Estresse Oxidativo/efeitos dos fármacos , 3-Hidroxiacil-CoA Desidrogenases/genética , 3-Hidroxiacil-CoA Desidrogenases/metabolismo , Acetil-CoA C-Aciltransferase/genética , Acetil-CoA C-Aciltransferase/metabolismo , Idoso , Aldeídos/administração & dosagem , Antioxidantes/administração & dosagem , Biomarcadores/sangue , Isomerases de Ligação Dupla Carbono-Carbono/genética , Isomerases de Ligação Dupla Carbono-Carbono/metabolismo , Carotenoides/metabolismo , Dieta/efeitos adversos , Enoil-CoA Hidratase/genética , Enoil-CoA Hidratase/metabolismo , Ácidos Graxos Ômega-3/metabolismo , Ácidos Graxos Ômega-6/metabolismo , Feminino , Humanos , Isoprostanos/administração & dosagem , Peroxidação de Lipídeos/efeitos dos fármacos , Peroxidação de Lipídeos/genética , Degeneração Macular/etiologia , Degeneração Macular/genética , Degeneração Macular/patologia , Masculino , Pessoa de Meia-Idade , Neuroprostanos/administração & dosagem , Oxirredução/efeitos dos fármacos , Estresse Oxidativo/genética , Racemases e Epimerases/genética , Racemases e Epimerases/metabolismo , Fatores de Risco
5.
Int J Mol Sci ; 20(16)2019 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-31434294

RESUMO

Our previous studies showed that microRNA-15a (miR-15a) was closely related to intramuscular fat (IMF) deposition in chickens; however, its regulatory mechanism remains unclear. Here, we evaluated the expression characteristics of miR-15a and its relationship with the expression of acetyl-CoA acyltransferase 1 (ACAA1), acyl-CoA oxidase 1 (ACOX1) and sterol carrier protein 2 (SCP2) by qPCR analysis in Gushi chicken breast muscle at 6, 14, 22, and 30 weeks old, where we performed transfection tests of miR-15a mimics in intramuscular preadipocytes and verified the target gene of miR-15a in chicken fibroblasts (DF1). The miR-15a expression level at 30 weeks increased 13.5, 4.5, and 2.7-fold compared with the expression levels at 6, 14, and 22 weeks, respectively. After 6 days of induction, miR-15a over-expression significantly promoted intramuscular adipogenic differentiation and increased cholesterol and triglyceride accumulation in adipocytes. Meanwhile, 48 h after transfection with miR-15a mimics, the expression levels of ACAA1, ACOX1 and SCP2 genes decreased by 56.52%, 31.18% and 37.14% at the mRNA level in intramuscular preadipocytes. In addition, the co-transfection of miR-15a mimics and ACAA1, ACOX1 and SCP2 3'UTR (untranslated region) dual-luciferase vector significantly inhibited dual-luciferase activity in DF1 cells. Taken together, our data demonstrate that miR-15a can reduce fatty acid oxidation by targeting ACAA1, ACOX1, and SCP2, which subsequently indirectly promotes the differentiation of chicken intramuscular preadipocytes.


Assuntos
Acetil-CoA C-Aciltransferase/metabolismo , Adipócitos/classificação , Adipócitos/metabolismo , Proteínas de Transporte/metabolismo , Diferenciação Celular/fisiologia , MicroRNAs/metabolismo , Acetil-CoA C-Aciltransferase/genética , Animais , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/metabolismo , Proteínas de Transporte/genética , Diferenciação Celular/genética , Galinhas , MicroRNAs/genética
6.
Hum Mutat ; 40(10): 1641-1663, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31268215

RESUMO

Mitochondrial acetoacetyl-CoA thiolase (T2, encoded by the ACAT1 gene) deficiency is an inherited disorder of ketone body and isoleucine metabolism. It typically manifests with episodic ketoacidosis. The presence of isoleucine-derived metabolites is the key marker for biochemical diagnosis. To date, 105 ACAT1 variants have been reported in 149 T2-deficient patients. The 56 disease-associated missense ACAT1 variants have been mapped onto the crystal structure of T2. Almost all these missense variants concern residues that are completely or partially buried in the T2 structure. Such variants are expected to cause T2 deficiency by having lower in vivo T2 activity because of lower folding efficiency and/or stability. Expression and activity data of 30 disease-associated missense ACAT1 variants have been measured by expressing them in human SV40-transformed fibroblasts. Only two variants (p.Cys126Ser and p.Tyr219His) appear to have equal stability as wild-type. For these variants, which are inactive, the side chains point into the active site. In patients with T2 deficiency, the genotype does not correlate with the clinical phenotype but exerts a considerable effect on the biochemical phenotype. This could be related to variable remaining residual T2 activity in vivo and has important clinical implications concerning disease management and newborn screening.


Assuntos
Acetil-CoA C-Acetiltransferase/genética , Acetil-CoA C-Aciltransferase/deficiência , Erros Inatos do Metabolismo dos Aminoácidos/genética , Predisposição Genética para Doença , Mutação , Acetil-CoA C-Acetiltransferase/química , Acetil-CoA C-Acetiltransferase/metabolismo , Acetil-CoA C-Aciltransferase/genética , Acetil-CoA C-Aciltransferase/metabolismo , Erros Inatos do Metabolismo dos Aminoácidos/diagnóstico , Erros Inatos do Metabolismo dos Aminoácidos/metabolismo , Animais , Regulação Enzimológica da Expressão Gênica , Estudos de Associação Genética , Variação Genética , Humanos , Redes e Vias Metabólicas , Modelos Moleculares , Fenótipo , Ligação Proteica , Conformação Proteica , Domínios e Motivos de Interação entre Proteínas , Multimerização Proteica , Relação Estrutura-Atividade
7.
Med J Malaysia ; 74(2): 174-175, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-31079130

RESUMO

Methylacetoacetyl-coenzyme A thiolase (MAT) deficiency is an autosomal recessive disease caused by a defect of mitochondrial acetoacetyl-CoA thiolase (T2). There is an error of isoleucine catabolism and ketone body utilization due to mutations in the acetyl-Coenzyme A acetyltransferase 1 (ACAT1) gene. We report a case of a 14 months old Sabahan boy with beta deficiency who presented with severe sepsis and ketoacidosis who subsequently recovered.


Assuntos
Acetil-CoA C-Aciltransferase/deficiência , Erros Inatos do Metabolismo dos Aminoácidos/diagnóstico , Acetil-CoA C-Acetiltransferase/genética , Acetil-CoA C-Acetiltransferase/metabolismo , Acetil-CoA C-Aciltransferase/genética , Adolescente , Erros Inatos do Metabolismo dos Aminoácidos/complicações , Erros Inatos do Metabolismo dos Aminoácidos/genética , Humanos , Isoleucina/metabolismo , Cetose/etiologia , Masculino , Sepse/etiologia
8.
Int J Cardiol ; 292: 218-224, 2019 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-31023563

RESUMO

BACKGROUND: Epicardial adipose tissue (EAT) is a risk factor for cardiovascular diseases. Glucagon-like peptide 1 analogs (GLP-1A) may have beneficial cardiovascular effects and reduce EAT, possibly throughout targeting GLP-1 receptor (GLP-1R). Nevertheless, the role of EAT GLP-1R, GLP-2R and their interplay with EAT genes involved in adipogenesis and fatty acid (FA) metabolism are unknown. We analyzed whether EAT transcriptome is related to GLP-1R/GLP-2R gene expression, and GLP-1/GLP-2 plasma levels in coronary artery disease patients (CAD). METHODS: EAT was collected from 17 CAD patients undergoing CABG for microarray analysis of GLP-1R, GLP-2R and genes involved in FA metabolism and adipogenesis. EAT thickness was measured by echocardiography. GLP-1 and GLP-2 levels were quantified by ELISA in CAD and healthy subjects (CTR). RESULTS: EAT GLP-1R was directly correlated with genes promoting beta-oxidation and white-to-brown adipocyte differentiation, and inversely with pro-adipogenic genes. GLP-2R was positively correlated with genes involved in adipogenesis and lipid synthesis, and inversely with genes promoting beta-oxidation. GLP-1 and GLP-2 levels were higher in CAD than CTR and in patients with greater EAT thickness. CONCLUSIONS: GLP-1 analogs may target EAT GLP-1R and therefore reduce local adipogenesis, improve fat utilization and induce brown fat differentiation. As EAT lies in direct contiguity to myocardium and coronary arteries, the beneficial effects of GLP-1 activation may extent to the heart. The increased levels of circulating GLP-1 and GLP-2 and EAT GLP-2R may be compensatory mechanisms related to CAD and also EAT expansion, but the meaning of these observations needs to be further investigated.


Assuntos
3-Hidroxiacil-CoA Desidrogenases/metabolismo , Acetil-CoA C-Aciltransferase/metabolismo , Tecido Adiposo Marrom/metabolismo , Tecido Adiposo Branco/metabolismo , Isomerases de Ligação Dupla Carbono-Carbono/metabolismo , Doenças Cardiovasculares/sangue , Enoil-CoA Hidratase/metabolismo , Receptor do Peptídeo Semelhante ao Glucagon 1/sangue , Pericárdio/metabolismo , Racemases e Epimerases/metabolismo , 3-Hidroxiacil-CoA Desidrogenases/genética , Acetil-CoA C-Aciltransferase/genética , Tecido Adiposo Marrom/diagnóstico por imagem , Tecido Adiposo Branco/diagnóstico por imagem , Adulto , Idoso , Antropometria/métodos , Isomerases de Ligação Dupla Carbono-Carbono/genética , Doenças Cardiovasculares/diagnóstico por imagem , Doenças Cardiovasculares/genética , Enoil-CoA Hidratase/genética , Feminino , Receptor do Peptídeo Semelhante ao Glucagon 1/genética , Humanos , Masculino , Pessoa de Meia-Idade , Pericárdio/diagnóstico por imagem , Racemases e Epimerases/genética , Fatores de Risco
10.
Nat Commun ; 9(1): 79, 2018 01 08.
Artigo em Inglês | MEDLINE | ID: mdl-29311546

RESUMO

Aromatic polyesters are widely used plastics currently produced from petroleum. Here we engineer Escherichia coli strains for the production of aromatic polyesters from glucose by one-step fermentation. When the Clostridium difficile isocaprenoyl-CoA:2-hydroxyisocaproate CoA-transferase (HadA) and evolved polyhydroxyalkanoate (PHA) synthase genes are overexpressed in a D-phenyllactate-producing strain, poly(52.3 mol% 3-hydroxybutyrate (3HB)-co-47.7 mol% D-phenyllactate) can be produced from glucose and sodium 3HB. Also, various poly(3HB-co-D-phenyllactate) polymers having 11.0, 15.8, 20.0, 70.8, and 84.5 mol% of D-phenyllactate are produced from glucose as a sole carbon source by additional expression of Ralstonia eutropha ß-ketothiolase (phaA) and reductase (phaB) genes. Fed-batch culture of this engineered strain produces 13.9 g l-1 of poly(61.9 mol% 3HB-co-38.1 mol% D-phenyllactate). Furthermore, different aromatic polyesters containing D-mandelate and D-3-hydroxy-3-phenylpropionate are produced from glucose when feeding the corresponding monomers. The engineered bacterial system will be useful for one-step fermentative production of aromatic polyesters from renewable resources.


Assuntos
Escherichia coli/metabolismo , Fermentação , Glucose/metabolismo , Engenharia Metabólica/métodos , Poliésteres/metabolismo , Ácido 3-Hidroxibutírico/metabolismo , Acetil-CoA C-Aciltransferase/genética , Acetil-CoA C-Aciltransferase/metabolismo , Aciltransferases/genética , Aciltransferases/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , /genética , Coenzima A-Transferases/genética , Coenzima A-Transferases/metabolismo , Cupriavidus necator/enzimologia , Cupriavidus necator/genética , Escherichia coli/genética , Lactatos/metabolismo , Oxirredutases/genética , Oxirredutases/metabolismo , Polietilenotereftalatos/metabolismo , Polímeros/metabolismo
11.
Sci Rep ; 8(1): 417, 2018 01 11.
Artigo em Inglês | MEDLINE | ID: mdl-29323178

RESUMO

Mammary epithelial cells (MECs) affect milk production capacity during lactation and are critical for the maintenance of tissue homeostasis. Our previous studies have revealed that the expression of miR-152 was increased significantly in MECs of cows with high milk production. In the present study, bioinformatics analysis identified ACAA2 and HSD17B12 as the potential targets of miR-152, which were further validated by dual-luciferase repoter assay. In addition, the expressions of miR-152 was shown to be negatively correlated with levels of mRNA and protein of ACAA2, HSD17B12 genes by qPCR and western bot analysis. Furthermore, transfection with miR-152 significantly up-regulated triglyceride production, promoted proliferation and inhibited apoptosis in MECs. Furthermore, overexpression of ACAA2 and HSD17B12 could inhibit triglyceride production, cells proliferation and induce apoptosis; but sh234-ACAA2-181/sh234-HSD17B12-474 could reverse the trend. These findings suggested that miR-152 could significantly influence triglyceride production and suppress apoptosis, possibly via the expression of target genes ACAA2 and HSD17B12.


Assuntos
17-Hidroxiesteroide Desidrogenases/genética , Acetil-CoA C-Aciltransferase/genética , Glândulas Mamárias Animais/citologia , MicroRNAs/genética , Triglicerídeos/metabolismo , 17-Hidroxiesteroide Desidrogenases/metabolismo , Regiões 3' não Traduzidas , Acetil-CoA C-Aciltransferase/metabolismo , Animais , Apoptose , Bovinos , Proliferação de Células , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Feminino , Regulação da Expressão Gênica , Lactação , Glândulas Mamárias Animais/metabolismo
12.
Curr Genet ; 64(2): 417-422, 2018 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-29043484

RESUMO

The precise and controlled regulation of gene expression at transcriptional and post-transcriptional levels is crucial for the eukaryotic cell survival and functions. In eukaryotes, more than 100 types of post-transcriptional RNA modifications have been identified. The N6-methyladenosine (m6A) modification in mRNA is among the most common post-transcriptional RNA modifications known in eukaryotic organisms, and the m6A RNA modification can regulate gene expression. The role of yeast m6A methyltransferase (Ime4) in meiosis, sporulation, triacylglycerol metabolism, vacuolar morphology, and mitochondrial functions has been reported. Stress triggers triacylglycerol accumulation as lipid droplets. Lipid droplets are physically connected to the different organelles such as endoplasmic reticulum, mitochondria, and peroxisomes. However, the physiological relevance of these physical interactions remains poorly understood. In yeast, peroxisome is the sole site of fatty acid ß-oxidation. The metabolic status of the cell readily governs the number and physiological function of peroxisomes. Under low-glucose or stationary-phase conditions, peroxisome biogenesis and proliferation increase in the cells. Therefore, we hypothesized a possible role of Ime4 in the peroxisomal functions. There is no report on the role of Ime4 in peroxisomal biology. Here, we report that IME4 gene deletion causes peroxisomal dysfunction under stationary-phase conditions in Saccharomyces cerevisiae; besides, the ime4Δ cells showed a significant decrease in the expression of the key genes involved in peroxisomal ß-oxidation compared to the wild-type cells. Therefore, identification and determination of the target genes of Ime4 that are directly involved in the peroxisomal biogenesis, morphology, and functions will pave the way to better understand the role of m6A methylation in peroxisomal biology.


Assuntos
Adenosina/análogos & derivados , Ácidos Graxos/genética , Metiltransferases/genética , Peroxissomos/genética , Proteínas de Saccharomyces cerevisiae/genética , 3-Hidroxiacil-CoA Desidrogenases/genética , Acetil-CoA C-Aciltransferase/genética , Adenosina/genética , Adenosina/metabolismo , Isomerases de Ligação Dupla Carbono-Carbono/genética , Enoil-CoA Hidratase/genética , Ácidos Graxos/metabolismo , Regulação Fúngica da Expressão Gênica/genética , Metabolismo dos Lipídeos/genética , Metiltransferases/metabolismo , Mitocôndrias/genética , Mitocôndrias/metabolismo , Peroxissomos/enzimologia , Processamento Pós-Transcricional do RNA/genética , Racemases e Epimerases/genética , Saccharomyces cerevisiae/genética , Vacúolos/enzimologia , Vacúolos/genética
13.
Metab Brain Dis ; 32(6): 2063-2071, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28875337

RESUMO

Hydroxysteroid (17ß) dehydrogenase 10 (HSD10) and mitochondrial acetoacetyl-CoA thiolase (ß-KT) are two adjacent enzymes for the degradation of isoleucine, thus HSD10 and ß-KT deficiencies are confusing at an early stage because of nearly the same elevation of typical metabolites in urine, such as 2-methyl-3-hydroxybutyric acid (2M3HBA) and tiglylglycine (TG). In order to better understand the differences between these two disorders, we described the clinical and molecular characteristics of two HSD10 deficiency patients and four ß-KT deficiency patients. ß-KT deficiency patients had a much more favorable outcome than that of HSD10 deficiency patients, indicating that the multifunction of HSD10, especially neurosteroid metabolic activity, other than only enzymatic degradation of isoleucine, is involved in the pathogenesis of HSD10 deficiency. Two different mutations, a novel mutation p.Ile175Met and a reported mutation p.Arg226Gln, were detected in the HSD17B10 gene of HSD10 deficiency patients. Six different mutations, including four known mutations: p.Ala333Pro, p.Thr297Lys, c.83_84delAT, c.1006-1G > C, and two novel mutations: p.Thr277Pro and c.121-3C > G were identified in the ACAT1 gene of ß-KT deficiency patients. In general, DNA diagnosis played an important role in distinguishing between these two disorders.


Assuntos
3-Hidroxiacil-CoA Desidrogenases/genética , Acetil-CoA C-Acetiltransferase/genética , Acetil-CoA C-Aciltransferase/deficiência , Erros Inatos do Metabolismo dos Aminoácidos/diagnóstico , Discinesias/diagnóstico , Epilepsia/genética , Isoleucina/metabolismo , Retardo Mental Ligado ao Cromossomo X/diagnóstico , Acetil-CoA C-Aciltransferase/genética , Acetil-CoA C-Aciltransferase/metabolismo , Erros Inatos do Metabolismo dos Aminoácidos/diagnóstico por imagem , Erros Inatos do Metabolismo dos Aminoácidos/genética , Erros Inatos do Metabolismo dos Aminoácidos/metabolismo , Encéfalo/diagnóstico por imagem , Pré-Escolar , China , Diagnóstico Diferencial , Discinesias/diagnóstico por imagem , Discinesias/genética , Discinesias/metabolismo , Epilepsia/metabolismo , Feminino , Humanos , Lactente , Masculino , Retardo Mental Ligado ao Cromossomo X/diagnóstico por imagem , Retardo Mental Ligado ao Cromossomo X/genética , Retardo Mental Ligado ao Cromossomo X/metabolismo , Modelos Moleculares , Mutação , Estudos Retrospectivos
14.
Appl Microbiol Biotechnol ; 101(21): 7945-7960, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28956111

RESUMO

The actinomycete Gordonia polyisoprenivorans strain VH2 is well-known for its ability to efficiently degrade and catabolize natural rubber [poly(cis-1,4-isoprene)]. Recently, a pathway for the catabolism of rubber by strain VH2 was postulated based on genomic data and the analysis of mutants (Hiessl et al. in Appl Environ Microbiol 78:2874-2887, 2012). To further elucidate the degradation pathway of poly(cis-1,4-isoprene), 2-dimensional-polyacrylamide gel electrophoresis was performed. The analysis of the identified protein spots by matrix-assisted laser desorption/ionization-time of flight tandem mass spectrometry confirmed the postulated intracellular pathway suggesting a degradation of rubber via ß-oxidation. In addition, other valuable information on rubber catabolism of G. polyisoprenivorans strain VH2 (e.g. oxidative stress response) was provided. Identified proteins, which were more abundant in cells grown with rubber than in cells grown with propionate, implied a putative long-chain acyl-CoA-dehydrogenase, a 3-ketoacyl-CoA-thiolase, and an aldehyde dehydrogenase. The amino acid sequence of the latter showed a high similarity towards geranial dehydrogenases. The expression of the corresponding gene was upregulated > 10-fold under poly(cis-1,4-isoprene)-degrading conditions. The putative geranial dehydrogenase and a homolog were purified and used for enzyme assays. Deletion mutants for five aldehyde dehydrogenases were generated, and growth with poly(cis-1,4-isoprene) was investigated. While none of the mutants had an altered phenotype regarding growth with poly(cis-1,4-isoprene) as sole carbon and energy source, purified aldehyde dehydrogenases were able to catalyze the oxidation of oligoisoprene aldehydes indicating an involvement in rubber degradation.


Assuntos
Aldeídos/metabolismo , Gordonia (Bactéria)/enzimologia , Gordonia (Bactéria)/metabolismo , Hemiterpenos/metabolismo , Látex/metabolismo , Oxirredutases/metabolismo , Acetil-CoA C-Aciltransferase/genética , Acetil-CoA C-Aciltransferase/metabolismo , Aldeído Desidrogenase/genética , Aldeído Desidrogenase/metabolismo , Carbono/metabolismo , Eletroforese em Gel Bidimensional , Metabolismo Energético , Deleção de Genes , Perfilação da Expressão Gênica , Gordonia (Bactéria)/genética , 3-Hidroxiacil-CoA Desidrogenase de Cadeia Longa/genética , 3-Hidroxiacil-CoA Desidrogenase de Cadeia Longa/metabolismo , Redes e Vias Metabólicas/genética , Oxirredução , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Espectrometria de Massas em Tandem
15.
Microb Cell Fact ; 16(1): 144, 2017 Aug 17.
Artigo em Inglês | MEDLINE | ID: mdl-28818103

RESUMO

BACKGROUND: In recent years the production of biobased biodegradable plastics has been of interest of researchers partly due to the accumulation of non-biodegradable plastics in the environment and to the opportunity for new applications. Commonly investigated are the polyhydroxyalkanoates (PHAs) poly(hydroxybutyrate) and poly(hydroxybutyrate-co-hydroxyvalerate) (PHB-V). The latter has the advantage of being tougher and less brittle. The production of these polymers in bacteria is well established but production in yeast may have some advantages, e.g. the ability to use a broad spectrum of industrial by-products as a carbon sources. RESULTS: In this study we increased the synthesis of PHB-V in the non-conventional yeast Arxula adeninivorans by stabilization of polymer accumulation via genetic modification and optimization of culture conditions. An A. adeninivorans strain with overexpressed PHA pathway genes for ß-ketothiolase, acetoacetyl-CoA reductase, PHAs synthase and the phasin gene was able to accumulate an unexpectedly high level of polymer. It was found that an optimized strain cultivated in a shaking incubator is able to produce up to 52.1% of the DCW of PHB-V (10.8 g L-1) with 12.3%mol of PHV fraction. Although further optimization of cultivation conditions in a fed-batch bioreactor led to lower polymer content (15.3% of the DCW of PHB-V), the PHV fraction and total polymer level increased to 23.1%mol and 11.6 g L-1 respectively. Additionally, analysis of the product revealed that the polymer has a very low average molecular mass and unexpected melting and glass transition temperatures. CONCLUSIONS: This study indicates a potential of use for the non-conventional yeast, A. adeninivorans, as an efficient producer of polyhydroxyalkanoates.


Assuntos
Poliésteres/metabolismo , Saccharomycetales/metabolismo , Acetil-CoA C-Aciltransferase/genética , Acetil-CoA C-Aciltransferase/metabolismo , Aciltransferases/genética , Aciltransferases/metabolismo , Oxirredutases do Álcool/genética , Oxirredutases do Álcool/metabolismo , Técnicas de Cultura Celular por Lotes , Reatores Biológicos , Proteínas Fúngicas/metabolismo , Cromatografia Gasosa-Espectrometria de Massas , Microscopia Eletrônica de Transmissão , Lectinas de Plantas/genética , Lectinas de Plantas/metabolismo , Plasmídeos/genética , Plasmídeos/metabolismo , Poliésteres/análise , Poliésteres/química , Saccharomycetales/enzimologia , Saccharomycetales/crescimento & desenvolvimento
16.
Mol Genet Metab ; 122(1-2): 67-75, 2017 09.
Artigo em Inglês | MEDLINE | ID: mdl-28689740

RESUMO

2-methylacetoacetyl-coenzyme A thiolase (MAT) deficiency, also known as beta-ketothiolase deficiency, is an inborn error of ketone body utilization and isoleucine catabolism. It is caused by mutations in the ACAT1 gene and may present with metabolic ketoacidosis. In order to obtain a more comprehensive view on this disease, we have collected clinical and biochemical data as well as information on ACAT1 mutations of 32 patients from 12 metabolic centers in five countries. Patients were between 23months and 27years old, more than half of them were offspring of a consanguineous union. 63% of the study participants presented with a metabolic decompensation while most others were identified via newborn screening or family studies. In symptomatic patients, age at manifestation ranged between 5months and 6.8years. Only 7% developed a major mental disability while the vast majority was cognitively normal. More than one third of the identified mutations in ACAT1 are intronic mutations which are expected to disturb splicing. We identified several novel mutations but, in agreement with previous reports, no clear genotype-phenotype correlation could be found. Our study underlines that the prognosis in MAT deficiency is good and MAT deficient individuals may remain asymptomatic, if diagnosed early and preventive measures are applied.


Assuntos
Acetil-CoA C-Aciltransferase/deficiência , Erros Inatos do Metabolismo dos Aminoácidos/complicações , Erros Inatos do Metabolismo dos Aminoácidos/genética , Ácidos Graxos/metabolismo , Isoleucina/metabolismo , Corpos Cetônicos/metabolismo , Acetil-CoA C-Acetiltransferase/genética , Acetil-CoA C-Aciltransferase/genética , Adolescente , Adulto , Erros Inatos do Metabolismo dos Aminoácidos/diagnóstico , Erros Inatos do Metabolismo dos Aminoácidos/fisiopatologia , Criança , Pré-Escolar , Consanguinidade , Feminino , Estudos de Associação Genética , Humanos , Lactente , Recém-Nascido , Masculino , Mutação , Triagem Neonatal , Prognóstico , Estudos Retrospectivos , Adulto Jovem
17.
J Dairy Sci ; 100(8): 6285-6297, 2017 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-28624287

RESUMO

The acetyl-CoA acyltransferase 2 (ACAA2) gene encodes an enzyme of the thiolase family that is involved in mitochondrial fatty acid elongation and degradation by catalyzing the last step of the respective ß-oxidation pathway. The increased energy needs for gluconeogenesis and triglyceride synthesis during lactation are met primarily by increased fatty acid oxidation. Therefore, the ACAA2 enzyme plays an important role in the supply of energy and carbon substrates for lactation and may thus affect milk production traits. This study investigated the association of the ACAA2 gene with important sheep traits and the putative functional involvement of this gene in dairy traits. A single nucleotide substitution, a T to C transition located in the 3' untranslated region of the ACAA2 gene, was used in mixed model association analysis with milk yield, milk protein yield and percentage, milk fat yield and percentage, and litter size at birth. The single nucleotide polymorphism was significantly associated with total lactation production and milk protein percentage, with respective additive effects of 6.81 ± 2.95 kg and -0.05 ± 0.02%. Additionally, a significant dominance effect of 0.46 ± 0.21 kg was detected for milk fat yield. Homozygous TT and heterozygous CT animals exhibited higher milk yield compared with homozygous CC animals, whereas the latter exhibited increased milk protein percentage. Expression analysis from age-, lactation-, and parity-matched female sheep showed that mRNA expression of the ACAA2 gene from TT animals was 2.8- and 11.8-fold higher in liver and mammary gland, respectively. In addition, by developing an allelic expression imbalance assay, it was estimated that the T allele was expressed at an average of 18% more compared with the C allele in the udder of randomly selected ewes. We demonstrated for the first time that the variants in the 3' untranslated region of the ovine ACAA2 gene are differentially expressed in homozygous ewes of each allele and exhibit allelic expression imbalance within heterozygotes in a tissue-specific manner, supporting the existence of cis-regulatory DNA variation in the ovine ACAA2 gene. This is the first study reporting differential allelic imbalance expression of a candidate gene associated with milk production traits in dairy sheep.


Assuntos
Regiões 3' não Traduzidas , Acetil-CoA C-Aciltransferase/genética , Lactação/genética , Ovinos/genética , Regiões 3' não Traduzidas/genética , Acetil-CoA C-Aciltransferase/química , Alelos , Animais , Feminino , Estudos de Associação Genética , Leite
18.
Mol Biosyst ; 13(8): 1504-1511, 2017 Jul 25.
Artigo em Inglês | MEDLINE | ID: mdl-28632266

RESUMO

Xuesaitong injection (XST), which mainly consists of Panax notoginseng saponins, has been widely used for treating cardio-cerebral vascular diseases. However, the underlying mechanisms of XST associated with its cardioprotective effects are still unclear. To identify the potential target proteins of XST, two-dimensional gel electrophoresis (2-DE)-based proteomics was utilized to analyze the protein profile of myocardium in rats with myocardial ischemia/reperfusion (I/R) injury. The differentially expressed proteins were identified by matrix assisted laser desorption/ionization time-of-flight mass spectrometry. It is interesting that XST can alter the expression of 7 proteins, including pyruvate dehydrogenase E1 alpha (PDHA1), hydroxyacyl-coenzyme A dehydrogenase (HADHA), peroxiredoxin 3 (PRX3), gamma-enolase, acetyl-coenzyme A acyltransferase 2 (ACAA2), etc. Functional analysis revealed that those proteins were chiefly related to cardiac energy metabolism and oxidative stress. The cardioprotective effects of XST were further validated in H9c2 cardiac muscle cells with hypoxia/reoxygenation injury. We found that XST can promote the activity of PDH, an important enzyme related to the TCA cycle, as well as increase the intracellular content of acetyl-CoA and ATP. Moreover, XST also attenuated intracellular MDA release in H2O2-injured cardiac cells. This is the first study on the proteomic expression of XST-treated myocardium with I/R injury to reveal that the cardioprotective effects of XST may be attributed to the PDH-mediated restoration of aerobic glucose oxidation.


Assuntos
Fármacos Cardiovasculares/farmacologia , Medicamentos de Ervas Chinesas/farmacologia , Metabolismo Energético/efeitos dos fármacos , Regulação da Expressão Gênica , Traumatismo por Reperfusão Miocárdica/tratamento farmacológico , Fármacos Neuroprotetores/farmacologia , Saponinas/farmacologia , Acetil-CoA C-Aciltransferase/genética , Acetil-CoA C-Aciltransferase/metabolismo , Animais , Linhagem Celular , Metabolismo Energético/genética , Perfilação da Expressão Gênica , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Masculino , Subunidade alfa da Proteína Mitocondrial Trifuncional/genética , Subunidade alfa da Proteína Mitocondrial Trifuncional/metabolismo , Traumatismo por Reperfusão Miocárdica/genética , Traumatismo por Reperfusão Miocárdica/metabolismo , Traumatismo por Reperfusão Miocárdica/patologia , Miocárdio/metabolismo , Miocárdio/patologia , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/patologia , Estresse Oxidativo , Fosfopiruvato Hidratase/genética , Fosfopiruvato Hidratase/metabolismo , Piruvato Desidrogenase (Lipoamida)/genética , Piruvato Desidrogenase (Lipoamida)/metabolismo , Ratos , Ratos Sprague-Dawley , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz
19.
Metab Eng ; 42: 33-42, 2017 07.
Artigo em Inglês | MEDLINE | ID: mdl-28550000

RESUMO

ß-Oxidation is the ubiquitous metabolic strategy to break down fatty acids. In the course of this four-step process, two carbon atoms are liberated per cycle from the fatty acid chain in the form of acetyl-CoA. However, typical ß-oxidative strategies are not restricted to monocarboxylic (fatty) acid degradation only, but can also be involved in the utilization of aromatic compounds, amino acids and dicarboxylic acids. Each enzymatic step of a typical ß-oxidation cycle is reversible, offering the possibility to also take advantage of reversed metabolic pathways for applied purposes. In such cases, 3-oxoacyl-CoA thiolases, which catalyze the final chain-shortening step in the catabolic direction, mediate the condensation of an acyl-CoA starter molecule with acetyl-CoA in the anabolic direction. Subsequently, the carbonyl-group at C3 is stepwise reduced and dehydrated yielding a chain-elongated product. In the last years, several ß-oxidation pathways have been studied in detail and reversal of these pathways already proved to be a promising strategy for the production of chemicals and polymer building blocks in several industrially relevant microorganisms. This review covers recent advancements in this field and discusses constraints and bottlenecks of this metabolic strategy in comparison to alternative production pathways.


Assuntos
Acetilcoenzima A/metabolismo , Acetil-CoA C-Aciltransferase/metabolismo , Bactérias/metabolismo , Proteínas de Bactérias/metabolismo , Plásticos Biodegradáveis/metabolismo , Acetilcoenzima A/genética , Acetil-CoA C-Aciltransferase/genética , Bactérias/genética , Proteínas de Bactérias/genética , Oxirredução
20.
J Inherit Metab Dis ; 40(3): 395-401, 2017 05.
Artigo em Inglês | MEDLINE | ID: mdl-28220263

RESUMO

Beta-ketothiolase (T2) deficiency is an inherited disease of isoleucine and ketone body metabolism caused by mutations in the ACAT1 gene. Between 2005 and 2016, a total of 41 patients with T2 deficiency were identified at a medical center in northern Vietnam, with an estimated incidence of one in 190,000 newborns. Most patients manifested ketoacidotic episodes of varying severity between 6 and 18 months of age. Remarkably, 28% of patients showed high blood glucose levels (up to 23.3 mmol/L). Ketoacidotic episodes recurred in 43% of patients. The age of onset, frequency of episodes, and identified genotype did not affect patient outcomes that were generally favorable, with the exception of seven cases (five died and two had neurological sequelae). Custom-tailored acute and follow-up management was critical for a positive clinical outcome. Two null mutations, c.622C>T (p.Arg208*) and c.1006-1G>C (p.Val336fs), accounted for 66% and 19% of all identified ACAT1 mutant alleles, respectively. Most patients showed characteristic biochemical abnormalities. A newborn screening program could be expected to have a high yield in Vietnam. Investigation findings of haplotypes linked to the most common ACAT1 mutation (c.622C>T) are consistent with an ancient common founder of mutation-bearing chromosomes belonging to the Kinh ethnic population. The direct management and long-term follow-up of a large number of T2-deficient patients enabled us to study the natural history of this rare disease.


Assuntos
Acetil-CoA C-Aciltransferase/deficiência , Erros Inatos do Metabolismo dos Aminoácidos/genética , Acetil-CoA C-Aciltransferase/genética , Alelos , Feminino , Haplótipos/genética , Humanos , Recém-Nascido , Masculino , Mutação/genética , Triagem Neonatal/métodos , Vietnã
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