الملخص
Aims@#The aim of this study was to cultivate the fungus Aspergillus tubingensis (MO503), which was isolated and identified from Algerian soil using submerged fermentation. The focus was on the production of lipase, achieved through utilizing a minimal medium from agro-food industries. Specifically, the study investigated the potential of three waste sources – olive-pomace, Pistacia lentiscus fruit remains and olive mill wastewater as substrates for enhancing lipase production. @*Methodology and results@#The three aforementioned wastes were chosen to ascertain their value and determine the most cost-effective option. Factorial analysis of variance (ANOVA) was employed to evaluate the waste with a significant effect on lipase production, as this enables determining differences among the various waste sources. Growth monitoring revealed a maximum lipase activity of 1030 ± 0,039 U at pH 5.4 for olive pomace. A series of biochemical purification techniques displayed a visible band on the polyacrylamide gel obtained through SDS-PAGE. Lipolytic activity was evidenced by zymography in the presence of olive oil. The antibacterial activities of purified lipase exhibited a high sensitivity against Gram-negative bacteria compared to Gram-positive bacteria. @*Conclusion, significance and impact of study@#In addition to its activity against Gram-negative bacteria, the lipid degradation facilitated by these lipases in olive oil offers promising applications in the textile and therapeutic industries.
الملخص
AIM: With building a proliferation model of PA-induced VSMC, the effect of ATGL, a key fat metabolism enzyme, on the phenotype transformation of VSMC was preliminarily explored. METHODS: 40 μmol/L Atglistatin was added to the proliferation model of VSMC induced by PA (50 μmol/L, 100 μmol/L, and 200 μmol/L, respectively) at separately administered concentrations, and cell viability and cell proliferation were detected by CCK-8 and EDU; cell migration ability was detected by scratch assay; oil red staining was used to detect the accumulation of lipid droplets in VSMC was detected by oil red staining; the effects of PA on ATGL as well as the effects of smooth muscle contraction phenotype proteins were examined by Western blot. RESULTS: PA at a concentration of 100 μmol/L could significantly induce VSMC proliferation, promote lipophagy and increase lipid droplet accumulation in VSMC; meanwhile, Atglistatin could exacerbate these changes caused by PA and increase lipid droplet accumulation in VSMC. CONCLUSION: Atglistatin exacerbates PA-induced VSMC proliferation and increases VSMC lipid droplet accumulation, and exacerbates transformation of proliferative phenotype of VSMC.
الملخص
Diacylglycerol (DAG) is an intermediate product in lipid metabolism and plays an important physiological role in human body. It is mainly prepared by hydrolyzing lipid with lipase. However, research on the detection method of 1, 2-diacylglycerol (1, 2-DAG) and 1, 3-diacylglycerol (1, 3-DAG) and catalytic specificity of lipase was not enough, which limits its wide application. To address these challenges, an efficient quantitative detection method was first established for 1, 2-DAG (0.025-0.200 g/L) and 1, 3-DAG (0.025-0.150 g/L) by combining supercritical fluid chromatography with evaporative light scattering detector and optimizing the detection and analysis parameters. Based on the molecular docking between Thermomyces lanuginosus lipase (TLL) and triolein, five potential substrate binding sites were selected for site-specific saturation mutation to construct a mutation library for enzyme activity and position specificity screening. The specificity of sn-1, 3 of the I202V mutant was the highest in the library, which was 11.7% higher than the specificity of the wild type TLL. In summary, the position specificity of TLL was modified based on a semi-rational design, and an efficient separation and detection method of DAG isomers was also established, which provided a reference for the study of the catalytic specificity of lipase.
الموضوعات
Humans , Diglycerides , Molecular Docking Simulation , Binding Sites , Catalysis , Lipase/geneticsالملخص
Monoacylglycerol lipase (MGL) is a serine hydrolase that plays a major role in the degradation of endogenous cannabinoid 2-arachidonoylglycerol. The role of MGL in some cancer cells has been confirmed, where inhibition of the MGL activity shows inhibition on cell proliferation. This makes MGL a promising drug target for the treatment of cancer. Recently, the development of covalent inhibitors of MGL has developed rapidly. These drugs have strong covalent binding ability, high affinity, long duration, low dose and low risk of drug resistance, so they have received increasing attention. This article introduces the structure and function of MGL, the characteristics, mechanisms and progress of covalent MGL inhibitors, providing reference for the development of novel covalent small molecule inhibitors of MGL.
الموضوعات
Monoacylglycerol Lipases/metabolism , Endocannabinoids/metabolismالملخص
This report presents a case of a male infant, aged 32 days, who was admitted to the hospital due to 2 days of bloody stools and 1 day of fever. Upon admission, venous blood samples were collected, which appeared pink. Blood biochemistry tests revealed elevated levels of triglycerides and total cholesterol. The familial whole genome sequencing revealed a compound heterozygous variation in the LPL gene, with one variation inherited from the father and the other from the mother. The patient was diagnosed with lipoprotein lipase deficiency-related hyperlipoproteinemia. Acute symptoms including bloody stools, fever, and bloody ascites led to the consideration of acute pancreatitis, and the treatment involved fasting, plasma exchange, and whole blood exchange. Following the definitive diagnosis based on the genetic results, the patient was given a low-fat diet and received treatment with fat-soluble vitamins and trace elements, as well as adjustments to the feeding plan. After a 4-week hospitalization, the patient's condition improved and he was discharged. Follow-up showed a decrease in triglycerides and total cholesterol levels. At the age of 1 year, the patient's growth and psychomotor development were normal. This article emphasizes the multidisciplinary diagnosis and treatment of familial hyperlipoproteinemia presenting with symptoms suggestive of acute pancreatitis, including bloody ascites, in the neonatal period.
الموضوعات
Humans , Infant , Male , Acute Disease , Ascites , Cholesterol , Hyperlipoproteinemia Type I/genetics , Hyperlipoproteinemias , Lipoprotein Lipase/genetics , Pancreatitis , Triglyceridesالملخص
Introduction: Carbohydrate and lipid digestive enzymes are instrumental in the absorbability of nutrients associated to diabetes and obesity. This study evaluated hydroethanolic extracts of Piper nigrum leaf and Morinda lucida stem bark for antioxidant capacity and enzymes (carbohydrate and lipid digestive) inhibition. Methods: Colorimetric assays determined enzyme (?-amylase, ?-glucosidase, lipase and cholesterol esterase) inhibition and antioxidant capacity (total phenolic (TPC) and flavonoid (TFC) content, radical scavenging activity (DPPH, ABTS), and ferric reducing antioxidant power (FRAP)) of hydroethanolic ethanolic extracts, ethyl acetate and hexane fractions. Results: At 1 mg/ml extracts of P nigrum and M lucida inhibited ?- amylase (9.82±1.05 - 36.63±0.69 %) and ?-glucosidase (22.47±0.34 - 67.77±0.58 %) activities. At 100 µg/ml extracts and fractions inhibited lipase (56.72±1.11 - 81.61±0.71 %) and cholesterol esterase (18.14 ±0.79 - 36.84±0.70 %) activities. IC50 for ?- amylase (2.20±0.02 - 7.8±1.42 mg/ml), ?-glucosidase (0.16±0.01 - 3.74±0.01 mg/ml), lipase (8.58±2.57 - 53.03±5.20 µg/ml) and cholesterol esterase (172.20±5.12 - 419.80±4.55 µg/ml) were registered. At 4 mg/ml, P. nigrum presented a higher TPC (153.78±8.31 - 354.63±6.33 mg/ml), TFC (21.65±1.14 -33.86±0.00 mg/ml) than M lucida TPC (10.21±0.11 - 169.89±6.54 mg/ml), TFC (ND - 87.32±6.14 mg/ml). P nigrum presented radical scavenging (DPPH and ABTS) activity with IC50 0.12±0.00 - 1.27±0.01 mg/ml compared to 1.31±0.02 - 3.44±0.12 mg/ml of M lucida. The FRAP IC50 values were better for P nigrum (3.38±0.14- 4.48±1.05 mg/ml) than M lucida (3.34±1.32 - 15.4±2.03 mg/ml). Conclusion: P nigrum presented better antioxidant capacity and more effective on lipid digestive enzymes while M lucida was more effective on carbohydrate digestive enzymes.
الملخص
Enzymes of the archaea living in extreme environments are resistant to the challenging conditions. Lipase is among the important enzymes used in the industry and agriculture. In this study, the extracellular lipase from extremely halophilic archaeon Halolamina sp. was characterized for the first time. Optimum temperature for the enzyme activity was determined as 70oC, optimum pH was 7.0, and the optimum salt concentration was 3.6 M. Additionally, more than 70% of the enzyme activity was remained between pH 3.0-10.0 for 48 h as well as incubation of the enzyme at 70oC for 30 min increased its activity for 44%, and no activity loss was observed after incubation at 80oC. Also, presence of the metals increased the enzyme activity up to 88%. The enzyme was highly resistant to the organic solvents acetone, methanol, and DMSO while strong inhibition was caused by n-butanol. Among the detergents, the enzyme kept its activity substantially in the presence of SDS; however, other detergents caused inhibition of the enzyme activity. This characterization study showed that the lipase from the haloarchaeon Halolamina sp. is highly stable at the wide ranges of temperature and pH values as well as in the presence of diverse inhibitors. This enzyme is promising to be used in biotechnological applications.
الموضوعات
Enzyme Stability , Halobacteriales , Archaea , Lipaseالملخص
Abstract Objective: Lysosomal acid lipase deficiency (LAL-D) is an underdiagnosed autosomal recessive disease with onset between the first years of life and adulthood. Early diagnosis is crucial for effective therapy and long-term survival. The objective of this article is to recognize warning signs among the clinical and laboratory characteristics of LAL-D in pediatric patients through a scope review. Sources: Electronic searches in the Embase, PubMed, Livivo, LILACS, Web of Science, Scopus, Google Scholar, Open Gray, and ProQuest Dissertations and Theses databases. The dataset included observational studies with clinical and laboratory characteristics of infants, children and adolescents diagnosed with lysosomal acid lipase deficiency by enzyme activity testing or analysis of mutations in the lysosomal acid lipase gene (LIPA). The reference selection process was performed in two stages. The references were selected by two authors, and the data were extracted in June 2020. Summary of the findings: The initial search returned 1593 studies, and the final selection included 108 studies from 30 countries encompassing 206 patients, including individuals with Wolman disease and cholesteryl ester storage disease (CESD). The most prevalent manifestations in both spectra of the disease were hepatomegaly, splenomegaly, anemia, dyslipidemia, and elevated transaminases. Conclusions: Vomiting, diarrhea, jaundice, and splenomegaly may be correlated, and may serve as a starting point for investigating LAL-D. Familial lymphohistiocytosis should be part of the differential diagnosis with LAL-D, and all patients undergoing upper gastrointestinal endoscopy should be submitted to intestinal biopsy.
الموضوعات
Humans , Infant , Child , Adolescent , Adult , Cholesterol Ester Storage Disease/diagnosis , Cholesterol Ester Storage Disease/genetics , Cholesterol Ester Storage Disease/drug therapy , Wolman Disease/diagnosis , Wolman Disease/genetics , Sterol Esterase/genetics , Sterol Esterase/therapeutic use , Hepatomegalyالملخص
Although herbal medicines(HMs)are widely used in the prevention and treatment of obesity and obesity-associated disorders,the key constituents exhibiting anti-obesity activity and their molecular mechanisms are poorly understood.Recently,we assessed the inhibitory potentials of several HMs against human pancreatic lipase(hPL,a key therapeutic target for human obesity),among which the root-extract of Rhodiola crenulata(ERC)showed the most potent anti-hPL activity.In this study,we adopted an integrated strategy,involving bioactivity-guided fractionation techniques,chemical profiling,and biochemical assays,to identify the key anti-hPL constituents in ERC.Nine ERC fractions(retention time=12.5-35 min),obtained using reverse-phase liquid chromatography,showed strong anti-hPL activity,while the major constituents in these bioactive fractions were subsequently identified using liquid chromatography-quadrupole time-of-flight mass spectrometry(LC-Q-TOF-MS/MS).Among the identified ERC constituents,1,2,3,4,6-penta-O-galloyl-β-D-glucopyranose(PGG)and catechin gallate(CG)showed the most potent anti-hPL activity,with pIC50 values of 7.59±0.03 and 7.68±0.23,respectively.Further investigations revealed that PGG and CG potently inhibited hPL in a non-competitive manner,with inhibition constant(Ki)values of 0.012 and 0.082 μM,respectively.Collectively,our integrative analyses enabled us to efficiently identify and characterize the key anti-obesity constituents in ERC,as well as to elucidate their anti-hPL mechanisms.These findings provide convincing evidence in support of the anti-obesity and lipid-lowering properties of ERC.
الملخص
Metabolic homeostasis requires dynamic catabolic and anabolic processes. Autophagy, an intracellular lysosomal degradative pathway, can rewire cellular metabolism linking catabolic to anabolic processes and thus sustain homeostasis. This is especially relevant in the liver, a key metabolic organ that governs body energy metabolism. Autophagy's role in hepatic energy regulation has just begun to emerge and autophagy seems to have a much broader impact than what has been appreciated in the field. Though classically known for selective or bulk degradation of cellular components or energy-dense macromolecules, emerging evidence indicates autophagy selectively regulates various signaling proteins to directly impact the expression levels of metabolic enzymes or their upstream regulators. Hence, we review three specific mechanisms by which autophagy can regulate metabolism: A) nutrient regeneration, B) quality control of organelles, and C) signaling protein regulation. The plasticity of the autophagic function is unraveling a new therapeutic approach. Thus, we will also discuss the potential translation of promising preclinical data on autophagy modulation into therapeutic strategies that can be used in the clinic to treat common metabolic disorders.
الملخص
@#Introduction: This study was conducted to investigate the in-vitro lipid-lowering properties of ‘Saba’ banana peel pectin (SBP) extracted using three methods for its possible use as a dietary fibre ingredient. Methods: Pectin from ‘Saba’ banana peels were extracted using acid extraction (citric acid), enzymatic extraction (cellulase), and microwave-assisted extraction. In-vitro lipid-lowering assays were performed using spectrophotometry for pancreatic lipase inhibition and cholesterol binding, while liquid chromatography was used for bile acid-binding capacity. Results: Results revealed that all SBPs were not able to inhibit pancreatic lipase activity. However, all SBPs can notably bind to cholesterol and bile acids, taurocholate, and glycocholate. Acid-extracted pectin had the highest binding capacity to cholesterol (51.36%–55.07%) and glycocholate (27.37%), whereas all SBPs were similarly bound to taurocholate. Conclusion: The results of this study showed that acidextracted SBPs can significantly bind to cholesterol and bile acids, glycocholate and taurocholate, thereby indicating a possible reduction in lipid metabolism.
الملخص
Proteus mirabilis lipase (PML) features tolerance to organic solvents and great potential for biodiesel synthesis. However, the thermal stability of the enzyme needs to be improved before it can be used industrially. Various computational design strategies are emerging methods for the modification of enzyme thermal stability. In this paper, the complementary algorithm-based ABACUS, PROSS, and FoldX were employed for positive selection of PML mutations, and their pairwise intersections were further subjected to negative selection by PSSM and GREMLIN to narrow the mutation library. Thereby, 18 potential single-point mutants were screened out. According to experimental verification, 7 mutants had melting temperature (Tm) improved, and the ΔTm of K208G and G206D was the highest, which was 3.75 ℃ and 3.21 ℃, respectively. Five mutants with activity higher than the wild type (WT) were selected for combination by greedy accumulation. Finally, the Tm of the five-point combination mutant M10 increased by 10.63 ℃, and the relative activity was 140% that of the WT. K208G and G206D exhibited certain epistasis during the combination, which made a major contribution to the improvement of the thermal stability of M10. Molecular dynamics simulation indicated that new forces were generated at and around the mutation sites, and the rearrangement of forces near G206D/K208G might stabilize the Ca2+ binding site which played a key role in the stabilization of PML. This study provides an efficient and user-friendly computational design scheme for the thermal stability modification of natural enzymes and lays a foundation for the modification of PML and the expansion of its industrial applications.
الموضوعات
Enzyme Stability , Lipase/chemistry , Molecular Dynamics Simulation , Proteus mirabilis/metabolism , Solvents/chemistryالملخص
Lipoprotein lipase (LPL) is a key enzyme in lipid and lipoprotein metabolism. LPL mainly hydrolyzes triglyceride-rich lipoproteins and provides free fatty acid (FFAs) for metabolic tissues. LPL acts as a molecular bridge between lipoproteins and cell surface lipoprotein receptors, facilitating lipoprotein uptake. Recent studies have shown that LPL is widely expressed in tissues. LPL has a variety of physiological functions, Lipoprotein lipase (LPL) is a key enzyme in lipid and lipoprotein metabolism. LPL mainly hydrolyzes triglyceride-rich lipoproteins and provides free fatty acid (FFAs) for metabolic tissues. LPL acts as a molecular bridge between lipoproteins and cell surface lipoprotein receptors, facilitating lipoprotein uptake. Recent studies have shown that LPL is widely expressed in tissues. LPL has a variety of physiological functions, which regulates lipid metabolism and energy balance in the brain. Besides, it is closely related to Alzheimer's disease. This paper mainly reviews the latest research progress of LPL in the nervous system and provides new targets for the treatment and prevention.
الملخص
Aims@#This study was aimed to express Meyerozyma guilliermondii strain RT lipase using Komagataella phaffii X-33 expression system and its biochemical characterization and analyse the predicted structure of the product.@*Methodology and results@#Meyerozyma guilliermondii strain RT obtained from the previous study was used as the source of RT lipase gene. Extracellular M. guilliermondii strain RT lipase expression has significantly been improved up to 56 U/mg at 24 h cultivation in Yeast extract-Peptone-Dextrose (YPD) medium containing (in w/v): 1% yeast extract, 2% peptone, 2% dextrose with 0.5% v/v methanol induction. Characterization of RT lipase showed optimum activity at 45 °C and pH 9. It exhibited stability in the alkaline pH range (8 to 10) and retained 50% of its residual activity at 30 °C for 30 min. Substrate specificity analysis revealed that it preferred short to medium-chain triacylglycerols (C2-C12) with the highest activity towards caprylic acid (C8). Pairwise alignment revealed three substitutions (S2L, S92L and S193L) present in non-CTG-clade hosts (K. phaffii). Homology modelling (YASARA) was used to predict the structures of RT lipase [wild type (wt) and recombinant (rc)]. Mutational analysis of the structures showed the differences in loops that might attribute to the reduction of the optimum temperature from 75 °C (wt) to 45 °C (rc).@*Conclusion, significance and impact of study@#RT lipase was successfully overexpressed extracellularly using K. phaffii expression system with 91.8-fold higher specific activity than the native host. The conceptual advances on the importance of codon optimization before expressing a protein from a CTG-clade species in a non-CTG-clade yeast have been highlighted and the effect of the rare codon usage in recombinant protein characteristics has been evident.
الموضوعات
Candidaالملخص
Eurytrema coelomaticum is a trematode reported in the pancreatic ducts of ruminants. It is conjectured that may cause disorders in the pancreas, as well as digestive and metabolic processes dependent on them. This study, determined if there is an impairment of exocrine pancreatic function, and correlated it with parasite burden. Pancreas, blood, and fecal samples were collected from 119 bovines at a abattoir. Stool samples were subjected to the gelatin and x-ray film digestion tests (to detect the presence of trypsin in feces). Using blood samples, the following biochemical tests were performed: amylase, lipase, glucose, fructosamine, cholesterol, triglycerides, total protein, albumin, and globulins. Analyses were correlated with pancreatic parasite burden. Cattle with a high parasitic load presented higher incidence of negative tests in both gelatin digestion and x-ray film digestion tests (P < 0.001) when compared to non-parasitized animals and those with a low parasitic load. Changes in those tests only occurred if the parasitemia was moderate or severe. The activity of the amylase and lipase enzymes was significantly higher in animals with low parasitemia (P < 0.05), compared to non-parasitized animals and with a high parasitic burden. In this study, in cases of high parasitemia, negative results were observed in both gelatin and x-ray film in the feces digestion tests. However, the low infection of E. coelomaticum, higher levels of serum amylase and lipase that also indicated loss of pancreatic exocrine functions were reported.
Eurytrema coelomaticum, um trematódeo de ductos pancreáticos de ruminantes. Conjectura-se que possa ocasionar transtornos nas funções pancreáticas, mais especificamente nos processos digestivos e metabólicos dependentes destas. Neste estudo, o objetivo foi determinar se há comprometimento da função pancreática exócrina, correlacionado-a a carga parasitária. Foram utilizados pâncreas e respectivas amostras de sangue e fezes de 119 bovinos. As amostras de fezes foram submetidas aos testes de digestão da gelatina em tubo e digestão de filme radiográfico, ambos para detecção de tripsina nas fezes. Foram realizados os seguintes exames bioquímicos em amostras de sangue: amilase, lipase, glicemia, frutosamina, colesterol, triglicerídeos, proteínas totais, albumina e globulinas. Após isto, as análises bioquímicas foram correlacionadas com a quantidade numérica de parasitas encontrados no pâncreas (post-mortem). Houve maior quantidade de testes negativos (digestão do filme radiográfico e prova de digestão da gelatina) nos animais com alta carga parasitária (P < 0.001), quando comparados aos animais não parasitados e com baixa carga parasitária. Portanto, os exames supracitados se alteram somente se a quantidade de parasitas for moderada ou severa. As atividades das enzimas amilase e lipase foram significativamente maiores nos animais que apresentavam baixa parasitemia (P < 0.05), em comparação com os animais com alta carga parasitária e não parasitados. Conclui-se que em quadros de alta parasitemia há alteração significativa nos testes de digestão nas fezes, e que em quadros de baixa parasitemia há alterações significativas nos valores de amilase e lipase séricas, ambos comprovando alterações pancreáticas importantes, de acordo com o quadro de parasitemia.
الموضوعات
Animals , Cattle , Exocrine Pancreatic Insufficiency/parasitology , Pancreatitis/parasitology , Trematode Infections/complications , Trematode Infections/veterinary , Amylases/blood , Lipase/blood , Trematoda , Parasite Load/veterinaryالموضوعات
Humans , Cholesterol Ester Storage Disease , Lipase , Fibrosis , Dyslipidemias , Fatty Liver , Hepatomegalyالملخص
Las variantes de ANGPTL3 con pérdida de función están asociadas con efectos beneficiosos sobre el metabolismo lipídico y de carbohidratos y con riesgo reducido de enfermedad coronaria. Los cambios beneficiosos en los parámetros lipídicos que se obtienen con la inhibición de ANGPTL3 junto con la reducción en aterosclerosis que se observa en modelos animales y en estudios epidemiológicos de genética humana hacen de ANGPTL3 un nuevo objetivo terapéutico para prevenir las enfermedades cardiovasculares. Dos estrategias novedosas han surgido para inhibir esta proteína: un anticuerpo monoclonal y un oligonucleótido antisentido, con capacidad para reducir tanto el colesterol como los triglicéridos plasmáticos en forma notoria. Aunque el horizonte es promisorio, todavía no sabemos si los efectos de una variante presente desde el comienzo de la vida serán reproducidos por la inhibición de esta proteína que se realiza más tarde en la vida a través de una intervención farmacológica. (AU)
Loss-of-function ANGPTL3 variants are associated with beneficial effects on carbohydrate and lipid metabolism, and reduced risk of coronary heart disease. The beneficial changes in lipid parameters obtained by ANGPTL3 inhibition together with atheroprotection observed in animal models and in epi-demiological studies of human genetics make ANGPTL3 a new therapeutic target to prevent cardiovascular diseases. Two novel strategies have emerged to inhibit this protein: a monoclonal antibody and an antisense oligonucleotide, with the ability to significantly lower plasma cholesterol and triglycerides. Although the horizon is promising, we still do not know if the effects of a variant present from the beginning of life will be reproduced by the inhibition of this protein that takes place later in life through a pharmacological intervention. (AU)
الموضوعات
Humans , Dyslipidemias/drug therapy , Angiopoietin-like Proteins/therapeutic use , Angiopoietin-like Proteins/pharmacology , Triglycerides/blood , Cardiovascular Diseases/prevention & control , Cholesterol/blood , Oligonucleotides, Antisense/pharmacology , Antibodies, Monoclonal/metabolismالملخص
BACKGROUND The extracellular expression of enzymes in a secretion host such as Bacillus subtilis is a useful strategy in reducing the cost of downstream processing of industrial enzymes. Here, we present the first report of the successful extracellular expression in Bacillus subtilis WB800 of Geobacillus stearothermophilus lipase (T1.2RQ), a novel industriallydesirable thermostable lipolytic enzyme which has an excellent hydrolytic and transesterification activity. Signal peptides of a-amylase, extracellular protease, and lipase A, as well as two different promoters, were used in the secretion and expression of lipase T1.2RQ. RESULTS Lipase activity assay using p-nitrophenyl laurate showed that all three signal peptides directed the secretion of lipase T1.2RQ into the extracellular medium. The signal peptide of lipase A, resulted in the highest extracellular yield of 5.6 U/ml, which corresponds to a 6-fold increase over the parent Bacillus subtilis WB800 strain. SDS-PAGE and zymogram analysis confirmed that lipase T1.2RQ was correctly processed and secreted in its original size of 44 kDa. A comparison of the expression levels of lipase T1.2RQ in rich medium and minimal media showed that the enzyme was better expressed in rich media, with up to an 8-fold higher yield over minimal media. An attempt to further increase the lipase expression level by promoter optimization showed that, contrary to expectation, the optimized promoter exhibited similar expression levels as the original one, suggesting the need for the optimization of downstream factors. CONCLUSIONS The successful extracellular secretion of lipase T1.2RQ in Bacillus subtilis represents a remarkable feat in the industrial-scale production of this enzyme
الموضوعات
Geobacillus stearothermophilus/metabolism , Geobacillus stearothermophilus/chemistry , Bacillus subtilis/metabolism , Bacillus subtilis/chemistry , Geobacillus stearothermophilus/isolation & purification , Geobacillus stearothermophilus/genetics , Bacillus subtilis/isolation & purification , Bacillus subtilis/genetics , Lipase/chemistryالملخص
Resumen La hipertrigliceridemia (HTG) es un problema que se presenta con frecuencia en la práctica clínica. Su prevalencia en adultos es cercana al 10%. El espectro varía desde una predisposición que resulta en HTG solo en presencia de sobrepeso considerable o consumo excesivo de alcohol hasta mutaciones graves muy raras que pueden conducir a HTG grave en la infancia, incluso en ausencia de factores adicionales, como en el síndrome de quilomicronemia familiar (FCS, familial chylomicronemia syndrome). Este es un trastorno autosómico recesivo poco frecuente del metabolismo del quilomicrón que causa una importante elevación de los triglicéridos (>10 mmol/885 mg/dl). Esta condición está asociada con un riesgo significativo de pancreatitis aguda recurrente. La aproximación diagnóstica se logra mediante la caracterización fenotípica, y el hallazgo de la alteración genética ayuda a dar un diagnóstico más preciso. Además, se ha propuesto una puntuación clínica para el diagnóstico de FCS, pero necesita más validación. Las opciones de tratamiento disponibles para reducir los triglicéridos, como los fibratos y los ácidos grasos omega-3, no son eficaces en los pacientes con FCS. Actualmente, el único tratamiento sigue siendo una dieta de por vida muy baja en grasas, que reduce la formación de quilomicrones. Finalmente, los inhibidores de la apolipoproteína C-III están en desarrollo y podrían constituir opciones de tratamiento para estos pacientes. Considerando lo anterior, el objetivo de este artículo es realizar una revisión general sobre la HTG grave, con énfasis en el FCS, basados en la literatura disponible más reciente.
Abstract Hypertriglyceridemia (HTG) is a problem that occurs frequently in clinical practice. Its prevalence in adults is close to 10% and it varies between regions. The spectrum ranges from a disposition that results in HTG only in the presence of considerable overweight and/or excessive alcohol consumption to very rare serious mutations that can lead to severe HTG in childhood, even in the absence of additional factors such as familial chylomicronemia syndrome (FCS). This is a rare autosomal recessive disorder of chylomicron metabolism that causes a severe elevation in triglyceride levels (>10 mmol/885 mg/dL). This condition is associated with a significant risk of recurrent acute pancreatitis. Because this is a genetic condition, the optimal diagnostic strategy remains the genetic test. In addition, a clinical score for the diagnosis of FCS has been proposed but it needs further validation. Available treatment options to lower triglycerides, such as fibrates or omega-3 fatty acids, are not effective in patients with FCS. Currently, the cornerstone of treatment remains a very low-fat, lifetime diet that reduces chylomicron formation. Finally, apolipoprotein C-3 inhibitors are under development and may eventually be treatment options for these patients. The objective of this article is to carry out a general review of severe HTG, with an emphasis on FCS and based on the most recent available literature.
الموضوعات
Chylomicrons , Pancreatitis , Hyperlipoproteinemia Type IV , Hyperlipoproteinemia Type Iالملخص
Several species of the Myrcia genus have been used in folk medicine to treat diabetes. Therefore, the aim of this work was to investigate the inhibitory activity of α-glucosidase and pancreatic lipase in the crude extract (EBF) and in the ethyl acetate fraction (FFA) of Myrcia hatschbachii, as well as to identify isolated phenolic compounds and to evaluate the antioxidant property and preliminary in vitro toxicity against Artemia salina. EBF (IC50: 3.21 µg/mL) and FFA (IC50: 1.14 µg/mL) showed inhibitory activity superior to acarbose (IC50: 193.65 µg/mL). In addition, they showed inhibitory effects of pancreatic lipase (IC50: 556.58 µg/mL for EBF and 532.68 µg/mL for FFA), antioxidant potential, absence of preliminary toxicity and presence of gallic andellagic acids in FFA. The relevant results in the inhibition of α-glucosidase and pancreatic lipase motivate new studies for the development of herbal medicines that assist in the treatment of diabetic patients.
Varias especies del género Myrcia se han utilizado en la medicina popular para tratar la diabetes. Por lo tanto, el objetivo de este trabajo fue investigar la actividad inhibitoria de la α-glucosidasa y la lipasa pancreática en el extracto crudo (EBF) y en la fracción de acetato de etilo (FFA) de Myrcia hatschbachii, así como identificar compuestos fenólicos aislados y evaluar la propiedad antioxidante y toxicidad in vitro preliminar contra Artemia salina. EBF (IC50: 3.21 µg/mL) y FFA (IC50: 1.14 µg/mL) mostraron una actividad inhibitoria superior a la acarbosa (IC50: 193.65 µg/mL). Además, mostraron efectos inhibitorios de la lipasa pancreática (IC50: 556.58 µg/mL para EBF y 532.68 µg/mL para FFA), potencial antioxidante, ausencia de toxicidad preliminar y presencia de ácidos gálico y elágico en FFA. Los resultados relevantes en la inhibición de la α-glucosidasa y la lipasa pancreática motivan nuevos estudios para el desarrollo de medicamentos a base de hierbas que ayudan en el tratamiento de pacientes diabéticos.