فهرست مطالب

Biomacromolecular Journal - Volume:4 Issue:2, 2018
  • Volume:4 Issue:2, 2018
  • تاریخ انتشار: 1397/08/23
  • تعداد عناوین: 6
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  • Zeinab Mohammadi *, Mahdi Alijanianzadeh, Rassoul Khalilzadeh, Sirus Khodadadi Pages 65-77
    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 chemotrapy induced neutropenia. So development of an effective and inexpensive method to achieve monoPEGylated G-CSF form is required. Here it is focused on the PEGylation reaction engineering and the subsequent purification processes that should be optimized to minimize the costs inherent to the additional steps.
    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 achieved a purity around of 99% for pegylated rhG-CSF. Assay by FTIR revealed that PEGylation correctly 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 developed an experimental method to purify monoPEGylated rhG-CSF which is simple, fast, and low cost method with high efficiency.
    Keywords: Pegylation, rhG-CSF, Filgrastim, Purification, Ion exchange chromatography
  • Roya Pournejati, Hamid Reza Karbalaei, Heidari * Pages 78-92
    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.
    Keywords: Bacterial secondary metabolite, Bioactive compounds, Persian Gulf, DNA binding agents
  • Arezou Ghahghaei *, Bahare Hosseini, Nassim Faridi Pages 93-104
    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.
    Keywords: Crowding agents, β-Casein, Amyloid, α-Lactalbumin
  • Maryam Sharifi, Azadeh Mohammadgholi *, Nastaran Asghari, Moghaddam Pages 105-113
    Eucalyptus Sargentii belongs to the Myrtaceae family of Australian trees. One of the species of eucalyptus has cultivated in Iran. Eucalyptus extraction is a promising plant with anti-cancer activity.
    Materials and Method
    In this study, Growth inhibitory effects of Eucalyptus extraction and doxorubicin were assessed by MTT assay. The effect of Eucalyptus extraction 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.
    Results
    Our data showed that Doxorubicin (1.5 µg/ml), Eucalyptus (20μg/ml) 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 decrased in 48 hours. In the flowcytometry (PI) test, the highest apoptosis was observed at a concentration of combination of Doxorubicin and Eucalyptus. During the cell cycle, the highest percentage of cells in the G1 / G0 phase was observed in the treatment of the cells with combination.
    Discussion
    According to the results combination of Doxorubicin and Eucalyptus extraction showed that higher level apoptosis in comparison with agent separately. Furthermore by the increasing of BAX mRNA expression indicated that pro- apoptotic function.
    Keywords: A549 lung adenocarcinoma cell line, Bax, Bcl-2, doxorubicin, Eucalyptus, KRAS
  • Monir Shalbafan, Gholamreza Rezaei Behbehani * Pages 114-117
    Human serum albumin (HSA) is one of the main endogenous vehicles for biodistribution of molecules by blood plasma. Association constants and thermodynamic parameters for the interaction of HSA with Methotrexate were studied by docking. Docking studies provide remarkable information on binding sites of drugs, estimating the binding energies of each drug conformation with corresponding scores and functions. Docking study suggests that Methotrexate is able to interact with HSA by means of hydrogen bonds with one lysine , two arginine ,one Asparagine and one Glutamine residues, whereas the carbonyl group is inserted in a hydrophobic pocket, close to Trp-214.The Estimated of Gibbs free energies( ΔG° (kcal/mol)) is equal to -12.6 for the best model. The negative values of ΔG° indicate a spontaneous process. The association constant value (Ka ≈ 109 L•mol−1) is favorable for its efficient biodistribution by blood plasma. Studies using molecular modeling were performed to analyze the main intermolecular interactions between the Methotrexat and the amino acid residues present in the subdomain IIA interaction cavity.
    Keywords: Human Serum Albumin, Methotrexate, docking, Thermodynamic parameters
  • Shahriar Saeidian * Pages 118-126
    Peroxidase (EC 1.11.1.7) is one of the key enzymes controlling plant growth, differentiation and development. Peroxidase from black blueberry (Morus Nigara) leaves and friuts was purified using ammonium sulfate salt precipitation and Anion-exchange chromatography on DEAE-sepharose. 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 isoenzyme 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 friuts in presence of guaiacol and H2O2 were optimum after incubation at 40 and 45°C, respectively. Temperature stability profile showed that PGPL had maximum stability at 60°C and retained 79% activity after incubation for 30 minutes. PGPF was 100% stable for 1 h at 45°C and retained 45% activity after 4 h of preincubation. PGPL seemed to have considerable thermostability, which can be favorable in industrial operations for traditional brewing and food processing. Vmax of PGPL and PGPF were 16.8 and 25.3 unit/mg protein, so showed km of 6.8 and 5.6 mM, respectively. NaN3 and kojic acid were potent inhibitors of peroxidase and ZnSO4 showed that has an activatory effect on peroxidase in Leaves and fruits fo blueberry.
    Keywords: Guaiacol, Peroxidase, Morus nigra, Temperature