Biochemistry
Milad Saffary; Zolfaghar Lotfi; Amir Arasteh
Abstract
Aloysia citrodora is very important in Iranian traditional medicine. The aim of this study was to produce silver nanoparticles with aqueous extract of A. citrodora and investigate its antioxidant and inhibitory effects on the amyloid nanofibers from BSA as model protein. For synthesis of silver nanoparticles, ...
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Aloysia citrodora is very important in Iranian traditional medicine. The aim of this study was to produce silver nanoparticles with aqueous extract of A. citrodora and investigate its antioxidant and inhibitory effects on the amyloid nanofibers from BSA as model protein. For synthesis of silver nanoparticles, aqueous A. citrodora extract was prepared and one mM silver nitrate was added to solution. To confirm the presence and determine the size of silver nanoparticles, UV-Vis spectroscopy and TEM were used. The antioxidant effect of the extract was evaluated by DPPH and the production of amyloid fibers was measured by Congored absorption method. The toxicity effect of nanoparticles on neuroblasts was investigated by MTT method and the inhibitory properties of the extract on amyloid production were investigated by TEM. Caryophyllene (7.66%), phytol (7.47%) and limonene (2.68%) were the most abundant compounds. TEM also confirmed spherical crystalline nanoparticles with a size between 30–70 nm. By increasing the concentration of the extract, the antioxidant properties of nanoparticles increased, so that 97.35% of DPPH free radicals were inhibited at a concentration of 1 mg/ml. The highest inhibition of amyloid fibrils production was observed at concentration of 2 mg/ml by 96%, and the highest level of cytotoxicity at a concentration of 2 and with an IC50 of 2.25 mg/ml. The results showed that nanoparticles from A. citrodora leaves with anti–amyloid, antioxidant and anticancer properties can be considered as a new bio–nanomaterial in reducing Alzheimer's and cancer complications.
Amir Arasteh; Zolfaghar Lotfi; Mohammad Fazilati; Habibollah Nazem
Volume 6, Issue 3 , March 2018, , Pages 11-25
Abstract
The urease enzyme (EC.3.5.1.5) is from hydrolase group that catalyzes urea hydrolysis to ammonia and carbon dioxide. This enzyme has various applications in nitrogen metabolism, vaccine preparation, urea diagnosis kits, drinking industries, and so on. In this study, amyloid nano-fibrils from bovine serum ...
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The urease enzyme (EC.3.5.1.5) is from hydrolase group that catalyzes urea hydrolysis to ammonia and carbon dioxide. This enzyme has various applications in nitrogen metabolism, vaccine preparation, urea diagnosis kits, drinking industries, and so on. In this study, amyloid nano-fibrils from bovine serum albumin were used as a new scaffold for immobilizing the urease enzyme. The production of amyloid nano–fibers has been optimized with three techniques of Congord specrophotometry, Spectrofluorimetry and Spectropolarimetry, and the resulting fibrils have been confirmed by electron microscopy images. Then the urease enzyme was immobilized on the amyloid fibrils using glutaraldehyde molecules via cross-linked bridges and their kinetic factors were compared with the free enzyme. The highest amount of amyloid fibrils was obtained after 48 hours incubation of bovine serum albumin at a concentration of 10 mg.ml-1 and 70 ºC in a citrate-phosphate buffer pH 4. The immobilized enzyme had more reusability and stability than the free form and showed a higher activity and a smaller Km. Optimum temperature was improved from 40 ºC to 70 ºC and optimum pH was also improved from 6–7 to 6–9 in immobilized enzyme. In conclusion, amyloid fibrils with different chemical groups have been suitable for immobilization of urea enzyme. Improvement of kinetic properties and stability of urease enzyme by immobilizing on amyloid fibers allows for the widespread use of this enzyme in the related industries.
Amir Arasteh; Zolfaghar Lotfi; Mohammad Fazilat; Habibollah Nazem
Volume 6, Issue 2 , December 2017, , Pages 127-137
Abstract
Amyloid fibrils are a class of protein nanoparticles in which native proteins are converted into dense fibers. The aggregation can be pathogenic or non-pathogenic. Recently, these fibrils have been considered for the production of Bio–nanomaterials due to their unique structure. In this study, ...
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Amyloid fibrils are a class of protein nanoparticles in which native proteins are converted into dense fibers. The aggregation can be pathogenic or non-pathogenic. Recently, these fibrils have been considered for the production of Bio–nanomaterials due to their unique structure. In this study, Bovine serum albumin was used as a model protein to optimize the fibrillation process. Concentrations of 2-10 mg/ml were prepared in buffer at different pH values of 3-7 and placed at 0-70 ºC for 0 to 48 hours, and the amounts of produced amyloid fibrils were analyzed by spectrophotometry, fluorimetry and circular dichroism. The spectra obtained from the Congo red absorption method were compared on the basis of the maximum wavelength and absorption at maximum wavelengths with a sample containing Congo red. The concentration of 10 mg/ml of protein, which was 48 hours in buffer at pH = 4 at 50 ºC, produced the highest amounts of amyloid. Optimum conditions were obtained by ThT fluorescence and circular dichroism at a concentration of 10 mg/ml, pH = 4, 70 ºC and 48 hours. The presence of the fibers was confirmed by transmission electron microscope images. The insoluble structure and dimensions of amyloid fibrils can be describe them as new Bio–nanomaterials. Optimizing the production of these structures allows them to be produced at higher scales.
