Biomacromolecular Journal
Year:2018 | Volume:4 | Issue:2|Papers Count:7

Background: PEGylation is a valuable strategy for enhancing the pharmacokinetic properties of recombinant methionyl human granulocyte colony-stimulating factor (rh-Met-G-CSF, filgrastim) which is used to treat chemotherapy induced neutropenia. So development of a cost-effective method is required to attain monoPEGylated G-CSF form. Here it is focused on the PEGylation reaction engineering and purification processes that should be optimized to reduce the costs. Methods: In this study methoxy polyethylene glycol propionaldehydes (mPEG-ALD) 20 kDa MW was utilized to produce monoPEGylated rhG-CSF. PEGylation reaction was carried out at 4 ° C and pH 4. 5 in the presence of sodium cyanoborohydride and three mPEG-ALD: protein molar ratios (3: 1, 5: 1 and 10: 1). The PEGylation reaction was monitored with SDS-PAGE. Subsequently, isolation of the monoPEGylated form was achieved by cation exchange chromatography (CEC) method. The PEG attachment site was assessed by FTIR and structural characteristics of purified products (unPEGylated and PEGylated rhG-CSF) were investigated by CD and intrinsic fluorescence techniques. Results: The results showed optimal yield of monoPEGylated protein (60%). Also a purity around of 99% achieved for pegylated rhG-CSF. Assay by FTIR disclosed that PEGylation precisely was performed in N-terminus of rhG-CSF. Structural analysis by CD and intrinsic fluorescence indicated that rhG-CSF maintained secondary structure after modification by PEGylation. Conclusion: Overall, this study has expanded a pilot method to purify monoPEGylated rhG-CSF which is simple, fast, and economical method with high efficiency.

Natural products have historically been considered as a rich source of therapeutic agents and play a pivotal role in the development of medicinal compounds. Among microorganisms, marine bacteria have been recognized as a rich, promising and untapped resource. In the present work, we focused on Gram-positive bacterial population of Iranian coastal area of the Persian Gulf and study on bioactive potentials of the organic extracts from the isolates. The disc diffusion, DNA-interaction analysis along with MTT assay, and red blood cell hemolysis analysis were performed to evaluate antibacterial, cytotoxicity and anti-inflammatory activities of the extracts. We succeed to identify 6, 39, 34 and 21 novel bacterial producers of the bioactive metabolite having antibacterial, DNA interaction ability, cytotoxic activity, and anti-inflammatory effects, respectively. Approximately 15. 4% of the isolates produced antibacterial agents against S. aureus and P. aeruginosa, meanwhile the Bacillus genus revealed the strongest potential. Moreover, a significant correlation was observed between DNA binding ability of the extracts and their cytotoxicity effect on cancerous cell lines; although the chemical nature of the solvent used effects on it. The data of anti-inflammatory study showed that some bacterial extractions are more potent than dexamethasone by more than 90% RBC hemolysis protection. Our results demonstrate that the Gram-positive bacterial population of the Persian Gulf can be considered as a novel rich source of bioactive compounds with valuable therapeutic impact.

Amyloid aggregation is produced from deposition of intermediately folded protein states. Chaperones are assistant molecules that prevent aggregation of protein approximately. In this study we evaluated the effect of different molecular crowding agents (dextran, ficoll and PEG) on chaperoning effect of β-casein in preventing of aggregation of α-lactalbumin. Our results show that dextran and PEG increase the rate of aggregation of α-lactalbumin while ficoll decrease the rate of this aggregation. β-Casein, as a molecular chaperone, prevented aggregation of α-lactalbumin which increased in the presence of ficoll, while its protection activity decreased in the presence of dextran and PEG. The decrease in protection activity of β-casein in the presence of dextran and PEG, is maybe because of enhance in nonspecific interaction between β-casein and α-lactalbumin and environment, or because of the effect of dextran and PEG on the rate of amyloid fibril formation of α-lactalbumin. On the other hand, the increase in chaperone activity of β-casein in the presence of ficoll could be due to the effect of ficoll on the aggregation of target protein and/or reducing the nonspecific interaction between protein and environment. In summary, our data suggest that crowding agents have different effect on the aggregation of α-lactalbumin as well as chaperone activity of β-casein.

Eucalyptus sargentii belongs to the Myrtaceae family of Australian trees. One of the species of Eucalyptus has cultivated in Iran. Eucalyptus extract is a promising natural source of anti-cancer molecules. In this study, Growth inhibitory effects of Eucalyptus extract and doxorubicin were assessed by MTT assay. The effect of Eucalyptus extract and doxorubicin on apoptosis in the lung adenocarcinoma A549 cell line was investigated by flow cytometry. Cell cycle analysis was evaluated by flow cytometry. Level of Gene expression of Bax, Bcl-2 and KRAS were determined by Real time PCR. Our data showed that Doxorubicin (1. 5 µ g ml-1 ), Eucalyptus (20 μ g ml-1 ) and their combination inhibited proliferation and induced apoptosis in A549. Level of mRNA expression of Bax was increased while expression of Bcl-2 and KRAS were decreased at 48 h. During flowcytometry (PI) test revealed that the highest apoptosis occured at a concentration of combination of Doxorubicin and Eucalyptus. Thrugh the cell cycle, the highest percentage of cells in the G0/G1 phase was observed in the treatment with combination of Doxorubicin and Eucalyptus. According to the results combination of Doxorubicin and Eucalyptus extract showed higher level apoptosis in comparison with agents separately. Furthermore, increase of BAX mRNA expression indicated pro-apoptotic function.

Human serum albumin (HSA) is one of the main endogenous vehicles for biodistribution of molecules by blood plasma. Association equilibrium constants and thermodynamic parameters for the interaction of HSA with Methotrexate were studied by docking. Docking study provides remarkable information on binding sites of HSA with drugs, estimating the binding parameters in some specific places on biomolecules. Docking study suggests that Methotrexate interacts with one lysine, two arginine, one Asparagine and one Glutamine residues, specifically. The Estimated of Gibbs free energies (Δ G° ) is equal to-52. 67 kJ mol-1 for the best model.

Peroxidase (EC 1. 11. 1. 7) is one of the key enzymes controlling plant growth, differentiation and development. Peroxidase from black mulberry (Morus Nigara) leaves and fruits was purified using ammonium sulfate salt precipitation and anion-exchange chromatography on DEAE-Sepharose column. The preparation gave an overall yield of 28. 2%, 80-fold purification for leaves and of 12. 4%, 97-fold purification for peroxidase of fruits. Specific activity of 15. 3 and 31. 1 U mg-1 protein for peroxidase of leaves and fruits were earned, respectively. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed that the purified enzymes were homogeneous and peroxidase of leaves have two isoenzymes with molecular weight of approximately 56 and 33 kDa, but peroxidase of fruits has a isoenzyme with molecular weight of approximately 55 kDa. Activity of peroxidase in leaves and fruits in the presence of guaiacol and H2O2 were optimum after incubation at 40 and 45 ° C, respectively. Temperature stability profile showed that the purified Guaiacol peroxidase of leaves had maximum stability at 60 ° C and retained 79% activity after incubation for 30 min. PGPF was 100% stable for 1  h at 45 ° C and retained 45% of its initial activity after 4  h of preincubation at this temperature. The purified Guaiacol peroxidase of leaves seemed to have considerable thermostability, which can be favorable in industrial operations for traditional brewing and food processing. Kinetic parameters assesment for the purified Guaiacol peroxidase of leaves and purified Guaiacol peroxidase of friuts showed Vmax of 16. 8 and 25. 3 U mg-1, and Km of 6. 8 and 5. 6 mM, respectively. NaN3 and kojic acid were potent inhibitors of peroxidase and ZnSO4 showed an activatory effect on peroxidase in leaves and fruits of black mulberry.

Parkinson’ s disease (PD), is the second most common neurodegenerative disease in the world. The pathogenesis of PD is associated with α-Synuclein (α S) fibrillation. Previous works have indicated that blocking α S aggregation could be an effective strategy for the treatment of PD. Carbon nanotubes (CNTs) have specific properties that make them potentially useful in biomedicine and biotechnology. CNTs can access the brain, but no investigation has been done to survey the effect of single walled carbon nanotubes (SWNTs) or multi walled carbon nanotubes (MWNTs) on α S fibrillation. Through the use of Thioflavin T (ThT) fluorescence spectroscopy, transmission electron microscopy (TEM) and MTT assay, we found for the first time that both type of CNTs can significantly inhibit α S aggregation and subsequently change its neurotoxicity. While a complete mechanistic understanding remains to be elucidated, these data indicate that CNTs may have high therapeutic potential for the use against neuropathological features of PD.