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The α-amino acid ester acyltransferase (SAET) from Sphingobacterium siyangensis is one of the enzymes with the highest catalytic ability for the biosynthesis of l-alanyl-l-glutamine (Ala-Gln) with unprotected l-alanine methylester and l-glutamine. To improve the catalytic performance of SAET, a one-step method was used to rapidly prepare the immobilized cells (SAET@ZIF-8) in the aqueous system. The engineered Escherichia coli (E. coli) expressing SAET was encapsulated into the imidazole framework structure of metal organic zeolite (ZIF-8). Subsequently, the obtained SAET@ZIF-8 was characterized, and the catalytic activity, reusability and storage stability were also investigated. Results showed that the morphology of the prepared SAET@ZIF-8 nanoparticles was basically the same as that of the standard ZIF-8 materials reported in literature, and the introduction of cells did not significantly change the morphology of ZIF-8. After repeated use for 7 times, SAET@ZIF-8 could still retain 67% of the initial catalytic activity. Maintained at room temperature for 4 days, 50% of the original catalytic activity of SAET@ZIF-8 could be retained, indicating that SAET@ZIF-8 has good stability for reuse and storage. When used in the biosynthesis of Ala-Gln, the final concentration of Ala-Gln reached 62.83 mmol/L (13.65 g/L) after 30 min, the yield reached 0.455 g/(L·min), and the conversion rate relative to glutamine was 62.83%. All these results suggested that the preparation of SAET@ZIF-8 is an efficient strategy for the biosynthesis of Ala-Gln.
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Escherichia coli/genética , Glutamina , Zeolitas/química , AminoácidosRESUMEN
Delivering a potential drug is a predominant challenge in medicinal chemistry.in this study, bio organic compounds of Cymbopogon citratus was screened by analysing physiochemical properties like solubility, permeability, efficacy, toxicity, and metabolic stability. The optimization of drug potential against virulent protein was calculated by using docking algorithm Autodock 4.2.3. Structure based ligand docking reveals that the compounds having better inhibition potential against virulent enzymes with insoluble and impermeable activities. The organic compounds of Cymbopogon citratus were screened using Lipinski rule of five and ADME/T prediction for drug likeliness. The structure based ligand docking was done between bioactive compounds of plant and virulent protein that cause diseases. The interaction was visualized using Discovery studio and was studies. The molecular docking of bioactive compounds resulted in better inhibition potential with controlled lipophilicity level, without causing toxicity that harms the natural habitat of humans. The compounds, 1,3,4-trimethyl -3cyclohexene-1-carboxaldehyde exhibit binding energy -4.70 Kcal/mol followed by β-myrcene – 4.35 Kcal/mol and Geraniol -4.35 Kcal/mol. Hence, structure based ligand docking and in silico ADME/T studies revealed that the compounds have better inhibition potential against Apolipoprotein by improving the prediction of drug compounds.
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The presence of fat, beyond physiological limits, in organs, other than the adipose tissue, like the liver, the skeletal muscle, the heart and the pancreas etc is called ectopic fat. It causes specific organ dysfunction in the tissues concerned. The importance of the ectopic fat is that it is connected to peripheral tissue insulin resistance, obesity, metabolic syndrome etc. Though the molecular mechanisms underlying the specific organ dysfunctions are understood, still grey areas exists as to the source ofthe ectopic fat and how it finds it’s way to the specific sites of the target organs (intra-myocellular in skeletal muscle, hepatocyte cytoplasm of liver,epicardial surface andcoronary arteries of heart etc.).The molecular mechanisms involving the actualectopic deposition fat, are not clear. This article focuses on some of the grey areas in the pathogenesis of the ectopic fat deposition, besides reviewing brieflythe factsalready known in the literature about ectopic fat deposition.
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Anthocephalus indicus or A. indicus (family Rubiaceae; Hindi name KADAMBA or KADAM) is one of such ayurvedic remedy that has been mentioned in many Indian medicinal literatures. This article discusses about the medicinal principles of Anthocephalus indicus. In this communication, we reviewed the phytochemistry of Anthocephalus cadamba and its application in the cure of various ailments like diabetes mellitus, diarrhoea, fever, inflammation, haemoptysis, cough, vomiting, wounds, ulcers, debility and antimicrobial activity. The major constituents of the plant are triterpenes, triterpenoid glycosides, flavanoids, saponins, indole alkaloids; cadambine, cadamine, isocadambine, isodihydrocadambine. This review discusses the investigations made by various workers related to chemical constituents, pharmacological action and toxicological studies of this plant since years till date.
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Objective To investigate the expression of hormone sensitive lipase (HSL) and carnitine acyl transferase-1 (CAT-1) in Zucker rats after sleeve gastrectomy and to discuss the weight loss mechanism.Methods 30 male Zucker rats aging 10 weeks were randomly divided into 3 groups:the operation group (10 rats),the sham operation group (10 rats) and the diet-pairing group (10 rats).The rats were decapitated to retrieve the retroperitoneal adipose,mRNA and protein expression of HSL and CAT-1 gene of were detected by RTPCR and Western blot.Results As for the operation group,the weight decreased significantly after the operation comparing to the other two groups ((250±5.8) g,(370±10.0) g,(310±9.6) g,P＜0.05).mRNA and protein expressions of HSL and CAT-1 gene were all significantly higher in the operation group (P＜0.05).Conclusion SG can up-regulate the expressions of lipolysis gene HSL and CAT-1 in adipose in Zucker rats,promoting fat hydrolysis and increasing the energy expenditure,which is one of the important mechanisms of weight loss.
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Nor-ursane triterpenes are formed by the reduction of one or several methyl groups on the ursane triterpene carbon skeleton. According to the position of vacant methyl, they can be divided into eight types, including 23-nor, 24-nor, 28-nor, 29-nor, 30-nor, 2-nor, 3-nor, and 3,24-nor. Moreover, nor-ursane triterpenes have several pharmacological effects such as anti-cancer, anti-inflammatory, anti-bacterial, inhibition of acyl cholesterol acyl transferase (ACAT) and so on. In this study, more than 30 relevant literatures of nor-ursane triterpenoid compounds at home and abroad were reviewed. The plant distribution, pharmacological activity and structural characteristics of nor-ursane triterpenes were studied. Therefore, this review may provide the basis for further study of this type of compound.
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Objective To investigate the role of Ginsenosides Rg1 for non-alcoholic fatty liver disease by β-oxidation.Methods 120 SD rats were randomly divided into control group(CON),model group(HFD),Ginsenosides Rg1low,medium and high dose group (GLD ,GMD and GHD) ,sodium deoxycholate of bear treatment group (PDT ) ,20 rats in each group .After 4 and 8 weeks treatment ,the rats were sacrificed ,Pathology of hepatic tissue was tested by HE staining ,and liver function ,lipid levels ,hepatic ac-yl-CoA synthetase (CoASH1) ,carnitine acyl transferase I(CATI) and acetyl coenzyme A oxidase 1 (ACOX1) mRNA and protein expression were tested .Results After 4 weeks of treatment ,the liver function tested by HE staining only improved in GHD group . After 8 weeks ,there′s a little fat particles aggregation in PDT and GLD groups ,but no infiltration of fat in GMD and GHD groups . After 4 weeks ,AST ,ALT and AKP ,CHOL ,TG and LDL-C levels were significantly lower in PDT ,GLD ,GMD and GHD groups compared with HFD group (P<0 .05) ,which were significant declined 8 weeks later .After 4 weeks ,HDL-C level in four groups was significantly increased ,then reached the normal level 8 weeks later .After 4 weeks ,CoASH1 ,CATI and ACOX1 expressions in hepatic tissue of four groups were significantly increased ,which improved more obviously after eight weeks .Conclusion Ginsen-oside Rg1 can improves nonalcoholic fatty liver phenotype by regulation of β-oxidation .
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Objective To clone and express Staphylococcus aureus drug resistance adenylyltransferase gene in E .coli BL21 ,and to make the foundation for its function research .Methods Primers were designed on the basis of adenylyltransferase gene in gen‐bank ,PCR was used to amplify adenylyltransferase gene using Staphylococcus aureus genomic DNA as template .The obtained PCR production was attatched with pGEX‐4t‐1(+ ) plasmid ,and transformed into E .coli BL21 (DE3) .The recombinant plasmid was di‐gested by double enzyme digestion and identified by gene sequence .The recombinant protein was induced to expression by IPTG and identified by Western‐blotting .Results Using Staphylococcus aureus genome as a template ,the target fragment about 800 bp was successful amplified .After enzyme‐cutting and DNA‐sequencing ,the target fragment showed that the ORF begin with ATG ,end with TAG ,783 bp in length ,the predicted isoelectric point and molecular weight were 7 .75 and 29 × 103 ,and it was homology 99%homology with the reported sequence gene in genbank .SDS‐PAGE and Western‐blot showed the molecular weight of recombinant fusion protein was about 55 × 103 .Conclusion Adenylyltransferase gene of Staphylococcus aureus was successfully cloned and ex‐pressed in E .coli as a fusion protein ,which makes the foundation for the research of its function .
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Objective: To study the expression of calcium-sensing receptor (CaR) in human macrophages and the effect of CaR on macrophages converting to foam cells with its mechanism. Methods: Human macrophages were treated with ox-LDL (100 mg/L) for 48 h to induce foam cell formation, the experimental foam cells were cultured in 3 groups for 48 h respectively. Blank control group, Foam cell + CaR agonists (Agonists) group and Foam cell + CaR inhibitor (Inhibitor) group. The CaR expression in macrophages was measured by immuno-fluorescence and Western blot analysis. The positive foam cell formation was detected by oil red O staining, the intracellular cholesterol metabolism in foam cell was observed by HPLC, the inlfammatory cytokine secretion in foam cell culture supernatant was examined by ELISA, the protein expressions of CD36 and acyl-coA: cholesterol acyl-transferase (ACAT-1) in foam cells was measured by Western blot analysis. Results: Immuno-lfuorescence and Western blot indicated CaR is expressed in human macrophages. Compared with Control group, Oil Red O staining and HPLC showed that Agonists group had less positive foam cell formation, decreased CE, FC , TC, and Inhibitor group had more positive foam cell formation, increased CE, FC, TC, allP<0.05; ELISA presented that Agonists group had decreased TNF-α, MIF, increased anti-inflammatory cytokine IL-10, and Inhibitor group had increased TNF-α, MIF, decreased IL-10, allP<0.05; Western blot analysis indicated that Agonists group had decreased protein expressions of CD36, ACAT-1 in foam cells, and Inhibitor group had increased protein expressions of CD36 and ACAT-1. Conclusion: CaR is expressed in human macrophages, the higher CaR expression may inhibit macrophages converting to foam cells.
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OBJECTIVE:To investigate the regulation effects of policosanol on lowering cholesterol and its enzymatic mechanism.METHODS:The rats were randomly assigned into control group,policosanol prevention group (4.0 mg?kg-1?d-1),policosanol low-dose,medium-dose and high-dose groups (4.0 mg?kg-1?d-1,6.0 mg?kg-1?d-1,8.0 mg?kg-1?d-1),lovastatin group (positive control) and hyperlipoidemia model group.The last five groups were induced hyperlipoidemia model for 4 weeks.Blood samples were collected after 6 weeks administration (i.g.).The levels of TC,TG,LDL-C and HDL-C in the serum were determined.Body weight and liver weight were measured and hepatic index was calculated.The activity of lecithin cholesterol acyl transferase (LCAT) in serum,hepatic lipoprotein lipase (LPL) and hepatic lipase (HL) were detected.RESULTS:Policosanol remarkably decreased the levels of TC (ranged from 39.1% to 43.3%) and LDL-C (ranged from 66.6% to 80.7%) in serum and hepatic index (ranged from 11.1% to 11.8%) (P
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AIM: Based on the finding of adipophilin expression with the increase in cellular cholesterol, the aim of the present study was to look for the active site of adipophilin in cellular cholesteryl metabolism. METHODS: Mouse peritoneal macrophages were incubated with 80 mg/L Ox-LDL (Ox-LDL group) or 80 mg/L Ox-LDL plus 1 mmol/L adipophilin antisense oligonucleotides (Ox-LDL+antisense group), respectively. At the various time points, the incubated cell samples were observed with adipophilin immunofluorescence staining, flow cytometric analysis and cellular cholesterol analysis. RESULTS: The Ox-LDL+antisense group cells contained significantly lower cholesteryl ester (19.9?1.9) mg/g (protein) than that of cells in Ox-LDL group (46.6?3.4) mg/g (protein) at 4 days. From 12 h, expression of adipophilin in Ox-LDL group increased more quickly than that of the cells in Ox-LDL+antisense group. At day 4, the level of adipophilin expression in Ox-LDL group was significantly higher than that in Ox-LDL+antisense group. During the observation, the amount of Ox-r[CL-3H] LDL taking up increased gradually in both groups, however, from day 1 the taking up amount in Ox-LDL+antisense group was less than that in Ox-LDL group. There was a statistical difference between the two groups from day 2 to day 4. From 6 h to day 2, the relative ACAT activity increased in both groups. The relative ACAT activity kept unchanged from day 2 to day 4 in the two groups. At day 2, the relative ACAT activity in Ox-LDL+antisense group was significantly lower than that in Ox-LDL group. Correlative analysis between activity of ACAT and adipophilin expression showed than R2 were 0.6176 and 0.8212 (P