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Engineering a thermostable iron superoxide dismutase based on manganese superoxide dismutase from Thermus thermophilus

Paper ID Volume ID Publish Year Pages File Format Full-Text
34235 45011 2016 9 PDF Available
Title
Engineering a thermostable iron superoxide dismutase based on manganese superoxide dismutase from Thermus thermophilus
Abstract

•Two thermostable Fe-SODs were obtained by single-site mutation of Mn-SOD.•Two mutants fold accurately and exhibit higher activity than the wild-type Mn-SOD.•Both mutants were stable to high temperature and alkaline pH.

Three residues His29, His84 and His171 were found to be the ligands of metal ions of one hyperthermostable Mn-SOD produced by Thermus thermophilus HB27 after multiple sequence alignment. Conversion of these residues might cause a change of metal-binding specificity. Thus, three mutants His29Ala, His84Ala, and His171Ala were prepared. The mutant types changed, and the metal-binding specificities of His29Ala and His171Ala altered from Mn to Fe. The alpha-helix contents of His29Ala and His171Ala were 66% and 59% respectively, suggesting that the mutants folded with a reasonable secondary structure. His29Ala showed an activity comparable to that of the wild type, and His171Ala exhibited a 39.6% higher activity than the wild type. These two mutants were thermostable from 30 to 50 °C after incubation for 30 min. The activity of His171Ala retained 51% at 70 °C compared with 36.7% for His29Ala. Both mutants retained >80% residual activity in the pH range 5.0–10.0 (except their pI of 8.0) after incubation at 4 °C for 30 min. This study provides a good foundation for accelerating the development of thermostable Fe-SOD for application in the fields of medicine and cosmetics.

Graphical abstractThe site-directed mutations of a thermostable superoxide dismutase (SOD) from Thermus thermophilus HB27 can change the metal binding specificity of SOD from Mn to Fe. Furthermore, both mutants can exhibit much higher activity and stability than the wild-type Mn-SOD.Figure optionsDownload full-size imageDownload as PowerPoint slide

Keywords
Manganese superoxide dismutase; Site-directed mutagenesis; Multiple sequence alignment; Megaprimer-PCR; Metal-binding specificity; Thermostable Fe-SOD
First Page Preview
Engineering a thermostable iron superoxide dismutase based on manganese superoxide dismutase from Thermus thermophilus
Publisher
Database: Elsevier - ScienceDirect
Journal: Process Biochemistry - Volume 51, Issue 1, January 2016, Pages 39–47
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering