Improving enzymatic activities and thermostability of a tri-functional enzyme with SOD, catalase and cell-permeable activities

Research output: Contribution to journalArticle

Abstract

Synergistic action of major antioxidant enzymes, e.g., superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) is known to be more effective than the action of any single enzyme. Recently, we have engineered a tri-functional enzyme, 6His-MnSOD-TAT/CAT-MnSOD (M-TAT/CM), with SOD, CAT and cell-permeable activities. The protein actively internalized into the cells and showed superior protection against oxidative stress-induced cell death over native enzymes fused with TAT. To improve its molecular size, enzymatic activity and stability, in this study, MnSOD portions of the engineered protein were replaced by CuZnSOD, which is the smallest and the most heat resistant SOD isoform. The newly engineered protein, CAT-CuZnSOD/6His-CuZnSOD-TAT (CS/S-TAT), had a 42% reduction in molecular size and an increase in SOD and CAT activities by 22% and 99%, respectively. After incubation at 70 °C for 10 min, the CS/S-TAT retained residual SOD activity up to 54% while SOD activity of the M-TAT/CM was completely abolished. Moreover, the protein exhibited a 5-fold improvement in half-life at 70 °C. Thus, this work provides insights into the design and synthesis of a smaller but much more stable multifunctional antioxidant enzyme with ability to enter mammalian cells for further application as protective/therapeutic agent against oxidative stress-related conditions.

Details

Authors
  • Piriya Luangwattananun
  • Warawan Eiamphungporn
  • Napat Songtawee
  • Leif Bülow
  • Chartchalerm Isarankura-Na-Ayudhya
  • Virapong Prachayasittikul
  • Sakda Yainoy
Organisations
External organisations
  • Mahidol University
Research areas and keywords

Subject classification (UKÄ) – MANDATORY

  • Medical Biotechnology

Keywords

  • Catalase, CuZnSOD, HIV-1 TAT, Protein engineering
Original languageEnglish
Pages (from-to)50-59
Number of pages10
JournalJournal of Biotechnology
Volume247
Publication statusPublished - 2017 Apr 10
Publication categoryResearch
Peer-reviewedYes