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Volume 7, Issue 4 (Nov 2019)                   Res Mol Med (RMM) 2019, 7(4): 7-16 | Back to browse issues page


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Zarkar N, Nasiri Khalili M A, Khodadadi S, Zeinoddini M, Ghodrati Siahmazgi M, Faramarzi N. Site-Specific PEGylation of Recombinant Immunotoxin DAB389IL-2: Structural and Functional Assessment. Res Mol Med (RMM). 2019; 7 (4) :7-16
URL: http://rmm.mazums.ac.ir/article-1-323-en.html
Abstract:   (2904 Views)
Background: DAB389IL-2 is considered a fusion immunotoxin and it is used for the CTCL therapy. DAB389IL-2 includes of two distinct portions; the catalytic domain of diphtheria toxin and IL-2. DAB389IL-2 duo to the presence of a free cysteine residue (Cys 513 in IL-2 part) is prone to unwanted intramolecular and intermolecular disulfide bonds formation and aggregation problems. Aggregation is considered as the most common physical instability. PEGylation is an effective approach to increase the stability and half-life of therapeutic proteins.
Materials and methods: In this study, the PEGylation of recombinant DAB389IL-2 was performed by mPEG-vinylsulfone, through partial denaturation condition at 4 0C for 24 h. To confirm the PEGylation reaction, SDS-PAGE and Dynamic Light Scattering (DLS) was used. The structure of DAB389IL-2 and PEGylated immunotoxin DAB389IL-2 was analyzed using the circular dichroism (CD) and fluorescence methods. Also, K562 cells line were treated with various concentrations of DAB389IL-2 and conjugated form. In the following, the nuclease activity of DAB389IL-2 and PEGylated form was determined. Results: The SDS-PAGE result confirmed the site-specific PEGylation of DAB389IL-2. Spectroscopy results exhibited that the PEGylation doesn’t affect the protein native structure. Also, cytotoxicity assay and nuclease activity test confirmed that PEGylated protein induces death in K562 cells line and DNA degradation respectively.  
Conclusion: It is concluded that the PEGylated immunotoxin DAB389IL-2 has a proper structure and function; thus, PEGylated immunotoxin requires more survey due its unique properties.
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Type of Study: Research | Subject: Biochemistry
Received: 2019/08/5 | Accepted: 2019/11/28 | Published: 2019/11/28

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