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1.
Arch Microbiol ; 204(6): 313, 2022 May 11.
Artigo em Inglês | MEDLINE | ID: mdl-35543769

RESUMO

Fruit and vegetable wastes create unhygienic conditions and pose a environmental pollution. The utilization of such wastes as carbon sources for production of enzyme with microbial intervention could be an ecofriendly and profitable approach, apart from diminishing the waste load. The present investigation focused on the feasibility of using mosambi (Citrus limetta) peel as substrate for multienzyme production (pectinase, cellulase and amylase) through microbial intervention. Fifteen fungi were isolated from organic waste and screened in vitro their potential of biodegradation of mosambi peel through enzymes production. The best performing isolate was selected and identified as Trichoderma asperellum NG-125 (accession number-MW287256). Conditions viz. temperature, pH, incubation time and nutrient addition were optimized for efficient enzymes production. The maximum enzyme activity (U ml-1 min-1) of pectinase (595.7 ± 2.47), cellulase (497.3 ± 2.06) and amylase (440.9 ± 1.44) were observed at pH 5.5, incubation temperature of 30 °C after 10 days of fermentation. Moreover, macro-nutrients such as ammonium sulfate (0.1%) and potassium-di-hydrogen-ortho-phosphate (0.01%) further also enhanced the production of enzymes. The SDS-PAGE analysis of purified pectinase, cellulase and amylase using showed molecular mass of 43, 66 and 33 kDa, respectively. The enzyme retention activity (ERA) of aforesaid enzymes was also tested with four different natural fiber matrices viz., bagasse, rice husk, paddy straw and wheat straw. Among these, the maximum ERA was observed on bagasse matrix (pectinase-56.35%, cellulose-77.68% and amylase 59.54%). Enzymatic juice clarification yield obtained with test enzyme was 75.8%, as compared to 80.5% of commercial enzyme. The result indicates that T. asperellum may be exploited as multifaceted biocatalysis.


Assuntos
Celulase , Hypocreales , Trichoderma , Amilases/metabolismo , Celulase/metabolismo , Fermentação , Hypocreales/metabolismo , Poligalacturonase/metabolismo , Trichoderma/química
2.
Nan Fang Yi Ke Da Xue Xue Bao ; 42(4): 561-567, 2022 Apr 20.
Artigo em Chinês | MEDLINE | ID: mdl-35527492

RESUMO

OBJECTIVE: To assess the effect of early abdominal puncture drainage (APD) on autophagy and Nrf-2/HO-1 pathway in rats with severe acute pancreatitis (SAP) and explore the possibile mechanism. METHODS: Thirty-two male SD rats were randomly divided into sham-operated (SO) group, SAP group with retrograde injection of 4% sodium taurocholate, APD group with insertion of a drainage tube into the lower right abdomen after SAP induction, and APD + ZnPP group with intraperitoneal injection of 30 mg/kg ZnPP 12 h before APD modeling. Blood samples were collected from the rats 12 h after modeling for analysis of amylase and lipase levels and serum inflammatory factors. The pathological changes of the pancreatic tissue were observed with HE staining. Oxidative stress in the pancreatic tissue was detected with colorimetry, and sub-organelle structure and autophagy in pancreatic acinar cells were observed by transmission electron microscopy. The expressions of autophagy-related proteins and Nrf-2/HO-1 pathway were detected using RT-PCR and Western blotting. RESULTS: Compared with those in SAP group, the rats with APD treatment showed significantly alleviated pathologies in the pancreas, reduced serum levels of lipase, amylase and inflammatory factors, lowered levels of oxidative stress, and activated expressions of Nrf-2/HO-1 pathway in the pancreas. The ameliorating effect of ADP was significantly inhibited by ZnPP treatment before modeling. APD obviously reversed mitochondrial and endoplasmic reticulum damages and p62 accumulation induced by SAP. CONCLUSION: APD treatment can suppress oxidative stress and repair impaired autophagy in rats with SAP by activating the Nrf-2/HO-1 pathway, thereby reducing the severity of SAP.


Assuntos
Pancreatite , Doença Aguda , Amilases/efeitos adversos , Amilases/metabolismo , Animais , Autofagia , Drenagem , Lipase/efeitos adversos , Lipase/metabolismo , Masculino , Pâncreas/patologia , Pancreatite/induzido quimicamente , Pancreatite/cirurgia , Punções , Ratos , Ratos Sprague-Dawley
3.
J Agric Food Chem ; 70(15): 4690-4700, 2022 Apr 20.
Artigo em Inglês | MEDLINE | ID: mdl-35404598

RESUMO

Trehalose is an important rare sugar that protects biomolecules against environmental stress. We herein introduce a dual enzyme cascade strategy that regulates the proportion of cargos and scaffolds, to maximize the benefits of enzyme immobilization. Based upon the self-assembling properties of the shell protein (EutM) from the ethanolamine utilization (Eut) bacterial microcompartment, we implemented the catalytic synthesis of trehalose from soluble starch with the coimmobilization of α-amylase and trehalose synthase. This strategy improved enzymatic cascade activity and operational stability. The cascade system enabled the efficient production of trehalose with a yield of ∼3.44 g/(L U), 1.5 times that of the free system. Moreover, its activity was maintained over 12 h, while the free system was almost completely inactivated after 4 h, demonstrating significantly enhanced thermostability. In conclusion, an attractive self-assembly coimmobilization platform was developed, which provides an effective biological process for the enzymatic synthesis of trehalose in vitro.


Assuntos
Glucosiltransferases , Trealose , Amilases/metabolismo , Biocatálise , Enzimas Imobilizadas/metabolismo , Glucosiltransferases/metabolismo , Peptídeos/metabolismo , Trealose/metabolismo
4.
Oxid Med Cell Longev ; 2022: 5559151, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35126816

RESUMO

BACKGROUND: The Caoguo-4 decoction, a classical Mongolian medicine formula, is widely used to treat spleen deficiency diarrhea (SDD) in Mongolian for decades. Previously, the Caoguo-4 decoction volatile oil has been confirmed to be effective in ameliorating symptoms of spleen deficiency diarrhea in an animal model. However, the underlying mechanism of the Caoguo-4 decoction volatile oil is yet to be established. The aim of the current study was to investigate the antidiarrheal effects and mechanism of the Caoguo-4 decoction volatile oil. METHOD: Wistar rats were randomly divided into 5 groups of 10 animals including control, model, positive, Caoguo-4 decoction, and Caoguo-4 decoction volatile oil groups (10 rats in each group). All the rats, besides those in the control group, were induced to develop SDD by a bitter-cold purgation method with Xiaochengqi decoction. The antidiarrheal effect of Caoguo-4 decoction volatile oil was evaluated by pathological section, serum D-xylose and AMS content, plasma MTL content, and gut microbiota analysis via 16S rRNA sequencing. RESULTS: The results showed that the developed SDD rat model (model group) had decreased food intake, increased weight loss, soft stool, and bad hair color. When compared with the control group, serum was significantly reduced serum D-xylose and AML but increased MTL levels in the model group (p < 0.05). However, after treatment with either the Caoguo-4 decoction (the decoction group) or Smecta (the positive group) or volatile oil from the Caoguo-4 decoction (the volatile oil group), a significant increase in the serum D-xylose levels was observed. Additionally, AML levels significantly increased in the positive and volatile oil groups, and MTL levels significantly decreased in the decoction and volatile oil groups, when compared with the model group (p < 0.05). The pathological changes of the intestinal mucosa showed that the structure of the epithelium in the villi of the small intestine was affected, deformed, and incomplete in the model group when compared with the control group. However, either the decoction group or the volatile oil group recovered the villous morphology. The results of OTU analysis and alpha diversity analysis of intestinal bacteria showed that the intestinal microbiota of the SDD model rats showed an obvious decrease in richness and diversity of intestinal microbiota. But the intervention treatment of decoction and volatile oil could significantly recover the richness and diversity of intestinal microbiota. CONCLUSION: The intestinal microbiota destroyed in SDD modelling could be significantly improved by the Caoguo-4 decoction volatile oils, which provides reference for clinical medication.


Assuntos
Medicamentos de Ervas Chinesas/farmacologia , Microbioma Gastrointestinal/efeitos dos fármacos , Óleos Voláteis/farmacologia , Amilases/metabolismo , Animais , Bacteroidetes/genética , Bacteroidetes/isolamento & purificação , Cianobactérias/genética , Cianobactérias/isolamento & purificação , Diarreia/tratamento farmacológico , Modelos Animais de Doenças , Medicamentos de Ervas Chinesas/uso terapêutico , Fezes/microbiologia , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/patologia , Masculino , Óleos Voláteis/uso terapêutico , RNA Ribossômico 16S/análise , RNA Ribossômico 16S/genética , RNA Ribossômico 16S/metabolismo , Ratos , Ratos Wistar , Baço/patologia , Xilose/sangue
5.
J Steroid Biochem Mol Biol ; 218: 106048, 2022 04.
Artigo em Inglês | MEDLINE | ID: mdl-34973376

RESUMO

The objective of this study was to investigate the effects of orchiectomy (ORX) and testosterone replacement therapy (TRT) on redox balance and function of salivary glands. Forty-five young adult male Wistar rats (3 months old) were either castrated bilaterally or underwent fictitious surgery (SHAM) and were subsequently distributed into 3 groups: SHAM, ORX, and TRT (castrated rats that received an intramuscular injection of testosterone cypionate 10 mg/kg/weekly). All treatments started 4 weeks after castration (4 months old) and lasted 4 weeks (5 months old). At the end of treatment, pilocarpine-induced salivary secretion was collected to analyze salivary flow rate and biochemistry composition, and the parotid (PG) and submandibular (SMG) glands were sampled for redox balance markers and histomorphometric analyses. ORX increased salivary flow rate, calcium, phosphate, and chloride, and decreased total protein and amylase, while not changing the salivary buffer capacity, pH, sodium, and potassium compared to SHAM. TRT restored all salivary parameters to SHAM values. ORX increased oxidative lipid and protein damage, total antioxidant capacity, and uric acid in both salivary glands compared to SHAM. Superoxide dismutase, catalase, and glutathione peroxidase activities were greater only in the SMG of the ORX group in relation to SHAM. ORX decreased duct and acini area, while increasing connective tissue in the PG. On the other hand, ORX reduced duct area and increased acini area in the SMG compared to SHAM. TRT restored the redox balance and histomorphometric parameters to close to SHAM values in both salivary glands. Orchiectomy-induced salivary gland dysfunction was characterized by an increase in the salivary flow rate and changes in the secretion of total protein, amylase, and electrolytes, which are key factors, considered important for maintaining oral health status. To sum up, orchiectomy impaired the redox balance of the salivary glands. Our results also showed that TRT reversed the oxidative damage, morphological alterations, and salivary gland dysfunction induced by orchiectomy. Therefore, these results suggest an important action of testosterone on the redox balance and secretory ability of salivary glands.


Assuntos
Orquiectomia , Testosterona , Amilases/metabolismo , Animais , Antioxidantes/metabolismo , Antioxidantes/farmacologia , Masculino , Oxirredução , Ratos , Ratos Wistar , Glândulas Salivares/metabolismo , Testosterona/metabolismo
6.
J Appl Microbiol ; 132(4): 2894-2905, 2022 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-35094446

RESUMO

AIMS: In eukaryotic cells, chromatin remodelling complexes are essential for the accessibility of transcription factors to the specific regulating regions of downstream genes. Here, we identified an actin-like protein PoARP9 in cellulase production strain Penicillium oxalicum 114-2, which was an essential member of SWI/SNF complex. To investigate the physiological function of PoARP9 in transcriptional regulation, the coding gene Poarp9 was deleted in P. oxalicum 114-2. METHODS AND RESULTS: The absence of PoARP9 affected the colony growth on medium with glucose, cellulose or starch as sole carbon source. Meanwhile, the expression levels of major cellulase genes were all upregulated in ΔPoarp9 under the cellulase-inducing condition. In addition, the expression levels of amylase transcription activator AmyR as well as two major amylase genes were also increased in ΔPoarp9. CONCLUSIONS: These results demonstrated that chromatin remodelling affects the development and expression of cellulase and amylase in P. oxalicum. And the SWI/SNF complex member PoARP9 plays essential roles in these processes. SIGNIFICANCE AND IMPACT OF THE STUDY: This study provided new insights into the regulation of cellulase and development in P. oxalicum. And the regulatory function of SWI/SNF complex member ARP9 towards cellulase and amylase expression in P. oxalicum was verified for the first time.


Assuntos
Celulase , Penicillium , Actinas/genética , Actinas/metabolismo , Amilases/genética , Amilases/metabolismo , Celulase/metabolismo , Regulação Fúngica da Expressão Gênica , Penicillium/metabolismo
7.
J Oleo Sci ; 71(2): 301-310, 2022 Feb 03.
Artigo em Inglês | MEDLINE | ID: mdl-35034939

RESUMO

The intake of moderate oils and fats is necessary to maintain the body's energy balance, and the fatty acid composition of different oils and fats varies in their nutrition and function. The study aimed to investigate the effects of lard and vegetable blend oil on gut microbiota, intestinal enzyme activities, and blood routine. Kunming mice were assigned to the three groups: (1) Control group (CK) was gavage administration with distilled water, (2) Plant oil group (ZWY) was gavage administration with edible vegetable blend oil, (3) Lard group (DWY) was gavage administration with lard. After 42 days, microbiological, digestive enzymes, and blood routine were performed. Compared with the CK group, Escherichia coli, Lactobacilli, and Bifidobacteria were significantly decreased (p < 0.05), the activities of protease, cellulase, amylase, and xylanase were markedly reduced (p < 0.05), the hemoglobin was significantly increased (p < 0.05) in the ZWY group and DWY groups, and the hematocrit was increased in the ZWY group (p < 0.05), while other routine blood indices were increased (p > 0.05). Compared to the ZWY group, the activity of cellulase and amylase were significantly increased (p < 0.05), the intestinal microorganism and the routine blood indexes had no significant difference in the DWY group. Lard and vegetable blend oil diet affected the composition of the intestinal microorganisms, and the functions of digestive enzymes. Meanwhile, the levels of digestive enzymes may be correlated with the intestinal microbiota.


Assuntos
Gorduras Insaturadas na Dieta/administração & dosagem , Gorduras Insaturadas na Dieta/farmacologia , Gorduras na Dieta/administração & dosagem , Gorduras na Dieta/farmacologia , Microbioma Gastrointestinal/efeitos dos fármacos , Hematócrito , Hemoglobinas , Intestinos/enzimologia , Óleos Vegetais/administração & dosagem , Óleos Vegetais/farmacologia , Amilases/metabolismo , Animais , Bifidobacterium , Celulase/metabolismo , Testes Diagnósticos de Rotina , Escherichia coli , Feminino , Testes Hematológicos , Lactobacillus , Masculino , Camundongos Endogâmicos , Peptídeo Hidrolases/metabolismo , Organismos Livres de Patógenos Específicos
8.
World J Microbiol Biotechnol ; 38(2): 30, 2022 Jan 06.
Artigo em Inglês | MEDLINE | ID: mdl-34989888

RESUMO

Enzymes are biocatalysts that are widely used in different industries and generate billions of dollars annually. With the advancement of biotechnology, new enzymatic sources are being evaluated, especially microbial ones, in order to find efficient producers. Endophytic fungi are promising sources of biomolecules; however, Amazonian species are still poorly studied as to their enzymatic production potential. In this sense, the production of hydrolases (amylases, lipases, cellulases and pectinases) was evaluated in endophytic fungi isolated from the leaves, roots and stems of açai palms (Euterpe precatoria). A qualitative test was carried out to detect the enzymatic synthesis in each isolate, and the most promising ones were cultivated using submerged fermentation. The enzyme extracts were quantified to determine those with the greatest activity. Cellulolytic and amylolytic extracts showed the highest enzymatic activities and were partially characterized. Among 50 isolates, 82.9% produced pectinase, 58.5% produced cellulase, 31.7% produced amylase, and 12.2% produced lipase. Penicillium sp. L3 was the best producer of amylase and Colletotrichum sp. S1 was the best producer of cellulase in liquid medium cultivation. The amylolytic extract showed the highest enzymatic activity at pH 8.0 and 45 °C, and the cellulolytic extract at pH 5.0 and 35 °C. The cellulase and amylase produced by the endophytes had their molecular masses estimated between 38 and 76 kDa. These results indicate that endophytic fungi from the açai palm can be used as a new source of hydrolytic enzymes, which can be applied in numerous biotechnological processes.


Assuntos
Endófitos/enzimologia , Endófitos/metabolismo , Euterpe/microbiologia , Fungos/enzimologia , Fungos/metabolismo , Amilases/metabolismo , Biotecnologia/métodos , Celulase/metabolismo , Celulases/metabolismo , Colletotrichum , Fungos/classificação , Hidrólise , Lipase/metabolismo , Penicillium , Peptídeo Hidrolases , Poligalacturonase/metabolismo
9.
Acta Diabetol ; 59(4): 473-479, 2022 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-34782929

RESUMO

AIMS: Alterations of the exocrine pancreas have been reported in type 1 diabetes, but their contribution to the pathogenesis of the disease is poorly understood. Here, we investigated markers of exocrine pancreas dysfunction in individuals at-risk of developing type 1 diabetes. METHODS: Serum P-amylase and lipase levels were assessed in samples obtained from healthy controls, patients with new onset type 1 diabetes, relatives participating to the TrialNet Pathway to Prevention who were, at blood collection, autoantibody negative or positive for a single autoantibody (low-risk individuals), and positive for multiple autoantibodies (high-risk individuals). Linear mixed models were adopted to estimate variation of pancreatic enzymes among the groups and to evaluate the influence of high-risk HLA genotypes and residual beta cell function on exocrine pancreas function. RESULTS: In adults, but not children, reduced levels of P-amylase and lipase were shown in at-risk individuals, including (for P-amylase levels only) those at low-risk, and in T1Dnew. Furthermore, while high-risk HLA genotypes negatively affected P-amylase levels in autoantibody negative adult individuals, fasting C-peptide levels did not correlate with pancreatic enzyme levels. CONCLUSIONS: Exocrine pancreas dysfunction precedes the onset of type 1 diabetes in adult at-risk individuals and may be unrelated to fasting C-peptide levels.


Assuntos
Diabetes Mellitus Tipo 1 , Pâncreas Exócrino , Adulto , Amilases/metabolismo , Autoanticorpos/metabolismo , Biomarcadores/metabolismo , Humanos , Pâncreas/metabolismo , Pâncreas Exócrino/metabolismo
10.
Biotechnol Appl Biochem ; 69(2): 650-659, 2022 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-33655550

RESUMO

By reducing the activation energy, enzymes accelerate the chemical reaction; therefore, they are good alternative for industrial catalysts. Amylase is a suitable enzyme as a catalyst for the chemical decomposition of starch. This enzyme is of great importance, and its production is highly profitable. α-Amylase is among the most important amylases produced naturally by animals, plants, and microorganisms. Still, the α-amylases produced by bacteria have a special place in industry and commerce. Moreover, a large volume of this enzyme can be produced by selecting an appropriate and optimized host to clone and express the α-amylase gene. The present study briefly reviews the structure, application, sources, and hosts used to produce recombinant α-amylase.


Assuntos
Amilases , alfa-Amilases , Amilases/genética , Amilases/metabolismo , Animais , Bactérias/metabolismo , Amido/metabolismo , alfa-Amilases/metabolismo
11.
Br J Nutr ; 127(3): 321-334, 2022 02 14.
Artigo em Inglês | MEDLINE | ID: mdl-33749571

RESUMO

A12-week experiment was conducted to evaluate the influences of thiamine ongrowth performance, and intestinal mitochondrial biogenesis and function of Megalobramaamblycephala fed a high-carbohydrate (HC) diet. Fish (24·73 (sem 0·45) g) were randomly assigned to one of four diets: two carbohydrate (CHO) levels (30 and 45 %) and two thiamine levels (0 and 1·5 mg/kg). HC diets significantly decreased DGC, GRMBW, FIMBW, intestinal activities of amylase, lipase, Na+, K+-ATPase, CK, complexes I, III and IV, intestinal ML, number of mitochondrial per field, ΔΨm, the P-AMPK: T-AMPK ratio, PGC-1ß protein expression as well as the transcriptions of AMPKα1, AMPKα2, PGC-1ß, mitochondrial transcription factor A, Opa-1, ND-1 and COX-1 and 2, while the opposite was true for ATP, AMP and reactive oxygen species, and the transcriptions of dynamin-related protein-1, fission-1 and mitochondrial fission factor. Dietarythiamine concentrations significantly increased DGC, GRMBW, intestinal activities of amylase, Na+, K+-ATPase, CK, complexes I and IV, intestinal ML, number of mitochondrial per field, ΔΨm, the P-AMPK:T-AMPK ratio, PGC-1ß protein expression as well as the transcriptions of AMPKα1, AMPKα2, PGC-1ß, Opa-1, ND-1, COX-1 and 2, SGLT-1 and GLUT-2. Furthermore, a significant interaction between dietary CHO and thiamine was observed in DGC, GRMBW, intestinal activities of amylase, CK, complexes I and IV, ΔΨm, the AMP:ATP ratio, the P-AMPK:T-AMPK ratio, PGC-1ß protein expression as well as the transcriptions of AMPKα1, AMPKα2, PGC-1ß, Opa-1, COX-1 and 2, SGLT-1 and GLUT-2. Overall, thiamine supplementation improved growth performance, and intestinal mitochondrial biogenesis and function of M. amblycephala fed HC diets.


Assuntos
Carboidratos da Dieta , Biogênese de Organelas , Proteínas Quinases Ativadas por AMP/metabolismo , Monofosfato de Adenosina , Adenosina Trifosfatases/metabolismo , Trifosfato de Adenosina/metabolismo , Amilases/metabolismo , Animais , Carboidratos da Dieta/metabolismo , Carboidratos da Dieta/farmacologia , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/metabolismo , Tiamina/farmacologia
12.
Crit Rev Biotechnol ; 42(3): 325-341, 2022 May.
Artigo em Inglês | MEDLINE | ID: mdl-34420464

RESUMO

Amylases hydrolyze starch to diverse products including dextrins and progressively smaller polymers of glucose units. Thermally stable amylases account for nearly 25% of the enzyme market. This review highlights the structural attributes of the α-amylases from thermophilic bacteria. Heterologous expression of amylases in suitable hosts is discussed in detail. Further, specific value maximization approaches, such as protein engineering and immobilization of the amylases are discussed in order to improve its suitability for varied applications on a commercial scale. The review also takes into account of the immobilization of the amylases on nanomaterials to increase the stability and reusability of the enzymes. The function-based metagenomics would provide opportunities for searching amylases with novel characteristics. The review is expected to explore novel amylases for future potential applications.


Assuntos
Amilases , alfa-Amilases , Amilases/metabolismo , Bactérias/metabolismo , Estabilidade Enzimática , Engenharia de Proteínas , Amido , alfa-Amilases/química , alfa-Amilases/metabolismo
13.
Carbohydr Polym ; 275: 118693, 2022 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-34742420

RESUMO

In recent years enzymatic treatment of maize has been utilized in the wet-milling process to increase the yield of extracted starch, proteins, and other constituents. One of the strategies to obtain this goal is to add enzymes that break down insoluble cell-wall polysaccharides which would otherwise entrap starch granules. Due to the high complexity of maize polysaccharides, this goal is not easily achieved and more knowledge about the substrate and enzyme performances is needed. To gather information of both enzyme performance and increase substrate understanding, a method was developed using mass spectrometry imaging (MSI) to analyze degradation products from polysaccharides following enzymatic treatment of the maize endosperm. Different enzymes were spotted onto cryosections of maize kernels which had been pre-treated with an amylase to remove starch. The cryosections were then incubated for 17 h. before mass spectrometry images were generated with a MALDI-MSI setup. The images showed varying degradation products for the different enzymes observed as pentose oligosaccharides differing with regards to sidechains and the number of linked pentoses. The method proved suitable for identifying the reaction products formed after reaction with different xylanases and arabinofuranosidases and for characterization of the complex arabinoxylan substrate in the maize kernel. HYPOTHESES: Mass spectrometry imaging can be a useful analytical tool for obtaining information of polysaccharide constituents and enzyme performance from maize samples.


Assuntos
Oligossacarídeos/química , Zea mays/química , Amilases/metabolismo , Parede Celular/química , Endosperma/química , Endosperma/metabolismo , Espectrometria de Massas/métodos , Oligossacarídeos/análise , Polissacarídeos/análise , Polissacarídeos/química , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Amido/química , Xilanos/química , Zea mays/metabolismo
14.
Food Res Int ; 150(Pt A): 110735, 2021 12.
Artigo em Inglês | MEDLINE | ID: mdl-34865754

RESUMO

Maltopentaose-forming amylases are promising enzymes for their ability to hydrolyze starch and produce functional maltooligosaccharides. Two maltopentaose-forming amylase genes from Bacillus megaterium (BmMFA) and Saccharophagus degradans (SdMFA) were expressed heterologously and their characteristics were analyzed. BmMFA has substantial thermostability and SdMFA owns superior product specificity. The carbohydrate-binding module of SdMFA was fused with BmMFA and the fused protein showed ideal enzymatic properties and displayed potential for industrial production of maltopentaose. Under the optimized conditions, the final product containing 47.41% maltopentaose was obtained with a conversion rate of 92.67% from starch. This study provides a novel strategy for the directed modification of MFAses through protein fusion approach.


Assuntos
Amilases , Oligossacarídeos , Amilases/metabolismo , Especificidade por Substrato
15.
Pancreas ; 50(9): 1281-1286, 2021 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-34860812

RESUMO

OBJECTIVES: To evaluate the safety of irreversible electroporation (IRE) on swine pancreatic tissue including its effects on peripancreatic vessels, bile ducts, and bowel. METHODS: Eighteen Yorkshire pigs underwent IRE ablation of the pancreas successfully and without clinical complications. Contrast-enhanced computed tomography angiography and laboratory studies before the IRE ablation with follow-up computed tomography angiography, laboratory testing, and pathological examination up to 4 weeks postablation were performed. RESULTS: In a subset of cases, anatomic peripancreatic vessel narrowing was seen by 1 week postablation, persisting at 4 weeks postablation, without apparent functional impairment of blood flow. Laboratory studies revealed elevated amylase and lipase at 24 hours post-IRE, suggestive of acute pancreatitis, which normalized by 4 weeks post-IRE. There was extensive pancreatic tissue damage 24 hours after IRE with infiltration of immune cells, which was gradually replaced by fibrotic tissue. Ductal regeneration without loss of pancreatic acinar tissue and glandular function was observed at 1 and 4 weeks postablation. CONCLUSIONS: In our study, we demonstrated and confirmed the safety and minimal complications of IRE ablation in the pancreas and its surrounding vital structures. These results show the potential of IRE as an alternative treatment modality in patients with pancreatic cancer, especially those with locally advanced disease.


Assuntos
Eletroporação/métodos , Modelos Animais , Pâncreas/patologia , Neoplasias Pancreáticas/terapia , Amilases/metabolismo , Animais , Angiografia por Tomografia Computadorizada , Feminino , Humanos , Lipase/metabolismo , Pâncreas/irrigação sanguínea , Pâncreas/diagnóstico por imagem , Neoplasias Pancreáticas/diagnóstico por imagem , Reprodutibilidade dos Testes , Suínos , Fatores de Tempo , Tomografia Computadorizada por Raios X
16.
BMC Microbiol ; 21(1): 344, 2021 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-34911440

RESUMO

BACKGROUND: Catabolite control protein A (CcpA) regulates the transcription of lactate dehydrogenase and pyruvate formate-lyase in Streptococcus bovis, but knowledge of its role in response to different pH is still limited. In this study, a ccpA-knockout strain of S. bovis S1 was constructed and then used to examine the effects of ccpA gene deletion on the growth and fermentation characteristics of S. bovis S1 at pH 5.5 or 6.5. RESULTS: There was a significant interaction between strain and pH for the maximum specific growth rate (µmax) and growth lag period (λ), which caused a lowest µmax and a longest λ in ccpA-knockout strain at pH 5.5. Deletion of ccpA decreased the concentration and molar percentage of lactic acid, while increased those of formic acid. Strains at pH 5.5 had decreased concentrations of lactic acid and formic acid compared to pH 6.5. The significant interaction between strain and pH caused the highest production of total organic acids and acetic acid in ccpA-knockout strain at pH 6.5. The activities of α-amylase and lactate dehydrogenase decreased in ccpA-knockout strain compared to the wild-type strain, and increased at pH 5.5 compared to pH 6.5. There was a significant interaction between strain and pH for the activity of acetate kinase, which was the highest in the ccpA-knockout strain at pH 6.5. The expression of pyruvate formate-lyase and acetate kinase was higher in the ccpA-knockout strain compared to wild-type strain. The lower pH improved the relative expression of pyruvate formate-lyase, while had no effect on the relative expression of acetate kinase. The strain × pH interaction was significant for the relative expression of lactate dehydrogenase and α-amylase, both of which were highest in the wild-type strain at pH 5.5 and lowest in the ccpA-knockout strain at pH 6.5. CONCLUSIONS: Overall, low pH inhibited the growth of S. bovis S1, but did not affect the fermentation pattern. CcpA regulated S. bovis S1 growth and organic acid fermentation pattern. Moreover, there seemed to be an interaction effect between pH and ccpA deletion on regulating the growth and organic acids production of S. bovis S1.


Assuntos
Proteínas de Bactérias/metabolismo , Proteínas Repressoras/metabolismo , Streptococcus bovis/crescimento & desenvolvimento , Streptococcus bovis/metabolismo , Acetato Quinase/genética , Acetato Quinase/metabolismo , Acetiltransferases/metabolismo , Amilases/genética , Amilases/metabolismo , Animais , Proteínas de Bactérias/genética , Ácidos Carboxílicos/metabolismo , Fermentação , Regulação Bacteriana da Expressão Gênica , Concentração de Íons de Hidrogênio , L-Lactato Desidrogenase/genética , L-Lactato Desidrogenase/metabolismo , Mutação , Proteínas Repressoras/genética , Ruminantes/microbiologia
17.
Int J Mol Sci ; 22(24)2021 Dec 14.
Artigo em Inglês | MEDLINE | ID: mdl-34948204

RESUMO

The inositol 1,4,5-triphosphate receptor-associated 2 (IRAG2) is also known as Jaw1 or lymphoid-restricted membrane protein (LRMP) and shares homology with the inositol 1,4,5-triphosphate receptor-associated cGMP kinase substrate 1 (IRAG1). IRAG1 interacts with inositol trisphosphate receptors (IP3 receptors /IP3R) via its coiled-coil domain and modulates Ca2+ release from intracellular stores. Due to the homology of IRAG1 and IRAG2, especially in its coiled-coil domain, it is possible that IRAG2 has similar interaction partners like IRAG1 and that IRAG2 also modulates intracellular Ca2+ signaling. In our study, we localized IRAG2 in pancreatic acinar cells of the exocrine pancreas, and we investigated the interaction of IRAG2 with IP3 receptors and its impact on intracellular Ca2+ signaling and exocrine pancreatic function, like amylase secretion. We detected the interaction of IRAG2 with different subtypes of IP3R and altered Ca2+ release in pancreatic acinar cells from mice lacking IRAG2. IRAG2 deficiency decreased basal levels of intracellular Ca2+, suggesting that IRAG2 leads to activation of IP3R under unstimulated basal conditions. Moreover, we observed that loss of IRAG2 impacts the secretion of amylase. Our data, therefore, suggest that IRAG2 modulates intracellular Ca2+ signaling, which regulates exocrine pancreatic function.


Assuntos
Células Acinares/metabolismo , Cálcio/metabolismo , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Inositol 1,4,5-Trifosfato/metabolismo , Proteínas de Membrana/metabolismo , Pâncreas Exócrino/metabolismo , Amilases/metabolismo , Animais , Sinalização do Cálcio/fisiologia , Feminino , Masculino , Camundongos , Camundongos Knockout
18.
Nature ; 600(7887): 110-115, 2021 12.
Artigo em Inglês | MEDLINE | ID: mdl-34819672

RESUMO

The human microbiome encodes a large repertoire of biochemical enzymes and pathways, most of which remain uncharacterized. Here, using a metagenomics-based search strategy, we discovered that bacterial members of the human gut and oral microbiome encode enzymes that selectively phosphorylate a clinically used antidiabetic drug, acarbose1,2, resulting in its inactivation. Acarbose is an inhibitor of both human and bacterial α-glucosidases3, limiting the ability of the target organism to metabolize complex carbohydrates. Using biochemical assays, X-ray crystallography and metagenomic analyses, we show that microbiome-derived acarbose kinases are specific for acarbose, provide their harbouring organism with a protective advantage against the activity of acarbose, and are widespread in the microbiomes of western and non-western human populations. These results provide an example of widespread microbiome resistance to a non-antibiotic drug, and suggest that acarbose resistance has disseminated in the human microbiome as a defensive strategy against a potential endogenous producer of a closely related molecule.


Assuntos
Acarbose/farmacologia , Farmacorresistência Bacteriana/efeitos dos fármacos , Microbioma Gastrointestinal/efeitos dos fármacos , Hipoglicemiantes/farmacologia , Inativação Metabólica , Metagenoma/genética , Boca/microbiologia , Fosfotransferases (Aceptor do Grupo Álcool)/genética , Acarbose/metabolismo , Amilases/metabolismo , Animais , Humanos , Hipoglicemiantes/metabolismo , Metagenoma/efeitos dos fármacos , Modelos Moleculares , Boca/efeitos dos fármacos , Fosfotransferases (Aceptor do Grupo Álcool)/química , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo
19.
Cells ; 10(11)2021 10 20.
Artigo em Inglês | MEDLINE | ID: mdl-34831029

RESUMO

Background: Although several approaches have revealed much about individual factors that regulate pancreatic development, we have yet to fully understand their complicated interplay during pancreas morphogenesis. Gfi1 is transcription factor specifically expressed in pancreatic acinar cells, whose role in pancreas cells fate identity and specification is still elusive. Methods: In order to gain further insight into the function of this factor in the pancreas, we generated animals deficient for Gfi1 specifically in the pancreas. Gfi1 conditional knockout animals were phenotypically characterized by immunohistochemistry, RT-qPCR, and RNA scope. To assess the role of Gfi1 in the pathogenesis of diabetes, we challenged Gfi1-deficient mice with two models of induced hyperglycemia: long-term high-fat/high-sugar feeding and streptozotocin injections. Results: Interestingly, mutant mice did not show any obvious deleterious phenotype. However, in depth analyses demonstrated a significant decrease in pancreatic amylase expression, leading to a diminution in intestinal carbohydrates processing and thus glucose absorption. In fact, Gfi1-deficient mice were found resistant to diet-induced hyperglycemia, appearing normoglycemic even after long-term high-fat/high-sugar diet. Another feature observed in mutant acinar cells was the misexpression of ghrelin, a hormone previously suggested to exhibit anti-apoptotic effects on ß-cells in vitro. Impressively, Gfi1 mutant mice were found to be resistant to the cytotoxic and diabetogenic effects of high-dose streptozotocin administrations, displaying a negligible loss of ß-cells and an imperturbable normoglycemia. Conclusions: Together, these results demonstrate that Gfi1 could turn to be extremely valuable for the development of new therapies and could thus open new research avenues in the context of diabetes research.


Assuntos
Proteínas de Ligação a DNA/deficiência , Diabetes Mellitus/metabolismo , Diabetes Mellitus/prevenção & controle , Fatores de Transcrição/deficiência , Células Acinares/citologia , Células Acinares/metabolismo , Amilases/metabolismo , Animais , Diferenciação Celular/genética , Proliferação de Células/genética , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Diabetes Mellitus/genética , Modelos Animais de Doenças , Regulação da Expressão Gênica , Grelina/metabolismo , Proteínas de Homeodomínio/metabolismo , Hiperglicemia/complicações , Hiperglicemia/genética , Integrases/metabolismo , Camundongos Transgênicos , Mutação/genética , Pâncreas/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
20.
Int J Immunopathol Pharmacol ; 35: 20587384211054036, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34696610

RESUMO

BACKGROUND AND OBJECTIVE: Living organisms respond to physical, chemical, and biological threats with a potent inflammatory response which alters organ cell signaling and leads to dysfunction. We evaluated the therapeutic effect of bone marrow-based mesenchymal stromal cell (BM-MSC) transplanted in rats to preserve tissue integrity and to restore homeostasis and function in the pancreatitis experimental pattern. METHODS: This study involved 40 adult male Wister rats. Repeated L-arginine injections caused chronic pancreatitis (CP), leading to the development of pancreatic damage and shifting the intracellular signaling pathways. Rats were then infused with BM-MSC labeled with PKH26 fluorescent linker dye for 12 weeks. RESULTS: Cell-surface indicators of BM-MSCs such as CD 90 and CD29 were expressed with the lack of CD34 expression. BM-MSC treatment considerably improved the alterations induced in a series of inflammatory markers, including IL-18, TNF-α, CRP, PGE2, and MCP-1. Furthermore, improvement was found in digestive enzymes and lipid profile with amelioration in myeloperoxidase activity. BM-MSC treatment also regulated the (TGF-/p-38MPAK/SMAD2/3) signaling factors that enhances repair of damaged pancreatic tissue, confirmed by reversed alteration of histopathological examination. CONCLUSION: our results further bring to light the promise of cell transplant therapy for chronic pancreatitis.


Assuntos
Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais , Pancreatite Crônica/terapia , Amilases/metabolismo , Animais , Arginina , Proteína C-Reativa/análise , Citocinas , Dinoprostona/sangue , Lipase/metabolismo , Metabolismo dos Lipídeos , Masculino , Pâncreas/enzimologia , Pâncreas/patologia , Pancreatite Crônica/sangue , Pancreatite Crônica/induzido quimicamente , Pancreatite Crônica/metabolismo , Ratos Wistar , Proteína Smad2/metabolismo , Proteína Smad3/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
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