TAT-BoNT/A(1᾿48), a novel fusion protein as a therapeutic agent: analysis of transcutaneous delivery and enzyme activity

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• 作者：Parvaneh Saffarian ; Shahin Najar Peerayeh…
• 关键词：Botulinum neurotoxin type A ; Protein purification ; TAT peptide ; Fusion protein ; Cell transduction ; Transcutaneous delivery
• 刊名：Applied Microbiology and Biotechnology
• 出版年：2016
• 出版时间：March 2016
• 年：2016
• 卷：100
• 期：6
• 页码：2785-2795
• 全文大小：1,393 KB
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• 作者单位：Parvaneh Saffarian (1)
  Shahin Najar Peerayeh (1)
  Jafar Amani (2)
  Firooz Ebrahimi (3)
  Hamid Sedighian (2)
  Raheleh Halabian (2)
  Abbas Ali Imani Fooladi (2)
  
  1. Department of Bacteriology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran
  2. Applied Microbiology Research Center, Baqiyatallah University of Medical Sciences, Tehran, Iran
  3. Department of Biology, Faculty of Basic Sciences, Imam Hussein University, Tehran, Iran
  
• 刊物类别：Chemistry and Materials Science
• 刊物主题：Chemistry
  Biotechnology
  Microbiology
  Microbial Genetics and Genomics
  
• 出版者：Springer Berlin / Heidelberg
• ISSN：1432-0614

文摘

Botulinum neurotoxin type A (BoNT/A) has been used as an injectable therapeutic agent for the treatment of some abnormal muscle contractions. In this study, TAT_(47–57) peptide, a cell-penetrating peptide, was fused with the catalytic domain of BoNT/A for therapeutic purposes. HeLa and BE(2)-C cell lines were treated separately with purified TAT-BoNT/A_(1–448) recombinant protein, and transduction of protein was analyzed by western blotting. Also, transcutaneous delivery through mouse skin surface was evaluated by immunohistochemistry. The in vitro catalytic activity of TAT-BoNT/A_(1–448) was evaluated by HPLC. The presence of recombinant protein was detected in both of the cell lines as well as mouse skin cryosections after 60 and 120 min of incubation. The concentration of intracellular proteins was increased over time. HPLC analysis showed that this fusion protein has a biological activity 1.5 times as much as the full-length BoNT/A_(1–448) protein. TAT-BoNT/A_(1–448) fusion protein is biologically active and can transmit through living cells in vitro and in vivo successfully and more effectively compared with BoNT/A_(1–448) protein as control.