A new gene encoding a thermostable Fe-superoxide dismutase (tcSOD) was identified from a metagenomic library prepared from a hot spring sample. The open reading frame of tcSOD encoded a 211 amino acid protein. The recombinant protein was overexpressed in Escherichia coli and confirmed to be a Fe–SOD with a specific activity of 1,890 U/mg using the pyrogallol method. The enzyme was highly stable at 80°C and retained 50% activity after heat treatment at 95°C for 2 h. It showed striking stability across a wide pH span from 4 to 11. The native form of the enzyme was determined as a homotetramer by analytical ultracentrifugation and gradient native polyacrylamide gel electrophoresis. Fe2+ was found to be important to SOD activity and to the stability of tcSOD dimer. Comparative modeling analyses of tcSOD tetramer indicate that its high thermostability is mainly due to the presence of a large number of intersubunit ion pairs and hydrogen bonds and to a decrease in solvent accessible hydrophobic surfaces.
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This work was supported by the National 863 High Technology Research and Development Program (grant no. 2004AA214080) and the National Natural Science Foundation of China (grant no. 30621005).
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He, Y., Fan, K., Jia, C. et al. Characterization of a hyperthermostable Fe-superoxide dismutase from hot spring. Appl Microbiol Biotechnol 75, 367–376 (2007). https://doi.org/10.1007/s00253-006-0834-3
- Sodium Dodecyl Sulfate
- Metagenomic Library
- Analytical Ultracentrifugation
- Subunit Interface
- Mesophilic Protein