Cloning and constitutive expression of <it>Deschampsia antarctica </it>Cu/Zn superoxide dismutase in <it>Pichia pastoris</it>
<p>Abstract</p> <p>Background</p> <p><it>Deschampsia antarctica </it>shows tolerance to extreme environmental factors such as low temperature, high light intensity and an increasing UV radiation as result of the Antarctic ozone layer thinning. It is very lik...
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doaj-874819a9f2734c7082466c9592e6a78a2020-11-25T01:46:20ZengBMCBMC Research Notes1756-05002009-10-012120710.1186/1756-0500-2-207Cloning and constitutive expression of <it>Deschampsia antarctica </it>Cu/Zn superoxide dismutase in <it>Pichia pastoris</it>Gidekel ManuelMoraga AnaBravo Ramírez León ADinamarca JorgeNavarrete AlejandroSánchez-Venegas Jaime R<p>Abstract</p> <p>Background</p> <p><it>Deschampsia antarctica </it>shows tolerance to extreme environmental factors such as low temperature, high light intensity and an increasing UV radiation as result of the Antarctic ozone layer thinning. It is very likely that the survival of this species is due to the expression of genes that enable it to tolerate high levels of oxidative stress. On that account, we planned to clone the <it>D. antarctica </it>Cu/ZnSOD gene into <it>Pichia pastoris </it>and to characterize the heterologous protein.</p> <p>Findings</p> <p>The Copper/Zinc superoxide dismutase (Cu/ZnSOD) gene, <it>SOD </it>gene, was isolated from a <it>D. antarctica </it>by cDNA library screening. This <it>SOD </it>gene was cloned in the expression vector pGAPZαA and successfully integrated into the genome of the yeast <it>P. pastoris </it>SMD1168H. A constitutive expression system for the expression of the recombinant SOD protein was used. The recombinant protein was secreted into the YPD culture medium as a glycosylated protein with a 32 mg/l expression yield. The purified recombinant protein possesses a specific activity of 440 U/mg.</p> <p>Conclusion</p> <p><it>D. antarctica </it>Cu/ZnSOD recombinant protein was expressed in a constitutive system, and purified in a single step by means of an affinity column. The recombinant SOD was secreted to the culture medium as a glycoprotein, corresponding to approximately 13% of the total secreted protein. The recombinant protein Cu/ZnSOD maintains 60% of its activity after incubation at 40°C for 30 minutes and it is stable (80% of activity) between -20°C and 20°C. The recombinant SOD described in this study can be used in various biotechnological applications.</p> http://www.biomedcentral.com/1756-0500/2/207 |
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DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Gidekel Manuel Moraga Ana Bravo Ramírez León A Dinamarca Jorge Navarrete Alejandro Sánchez-Venegas Jaime R |
spellingShingle |
Gidekel Manuel Moraga Ana Bravo Ramírez León A Dinamarca Jorge Navarrete Alejandro Sánchez-Venegas Jaime R Cloning and constitutive expression of <it>Deschampsia antarctica </it>Cu/Zn superoxide dismutase in <it>Pichia pastoris</it> BMC Research Notes |
author_facet |
Gidekel Manuel Moraga Ana Bravo Ramírez León A Dinamarca Jorge Navarrete Alejandro Sánchez-Venegas Jaime R |
author_sort |
Gidekel Manuel |
title |
Cloning and constitutive expression of <it>Deschampsia antarctica </it>Cu/Zn superoxide dismutase in <it>Pichia pastoris</it> |
title_short |
Cloning and constitutive expression of <it>Deschampsia antarctica </it>Cu/Zn superoxide dismutase in <it>Pichia pastoris</it> |
title_full |
Cloning and constitutive expression of <it>Deschampsia antarctica </it>Cu/Zn superoxide dismutase in <it>Pichia pastoris</it> |
title_fullStr |
Cloning and constitutive expression of <it>Deschampsia antarctica </it>Cu/Zn superoxide dismutase in <it>Pichia pastoris</it> |
title_full_unstemmed |
Cloning and constitutive expression of <it>Deschampsia antarctica </it>Cu/Zn superoxide dismutase in <it>Pichia pastoris</it> |
title_sort |
cloning and constitutive expression of <it>deschampsia antarctica </it>cu/zn superoxide dismutase in <it>pichia pastoris</it> |
publisher |
BMC |
series |
BMC Research Notes |
issn |
1756-0500 |
publishDate |
2009-10-01 |
description |
<p>Abstract</p> <p>Background</p> <p><it>Deschampsia antarctica </it>shows tolerance to extreme environmental factors such as low temperature, high light intensity and an increasing UV radiation as result of the Antarctic ozone layer thinning. It is very likely that the survival of this species is due to the expression of genes that enable it to tolerate high levels of oxidative stress. On that account, we planned to clone the <it>D. antarctica </it>Cu/ZnSOD gene into <it>Pichia pastoris </it>and to characterize the heterologous protein.</p> <p>Findings</p> <p>The Copper/Zinc superoxide dismutase (Cu/ZnSOD) gene, <it>SOD </it>gene, was isolated from a <it>D. antarctica </it>by cDNA library screening. This <it>SOD </it>gene was cloned in the expression vector pGAPZαA and successfully integrated into the genome of the yeast <it>P. pastoris </it>SMD1168H. A constitutive expression system for the expression of the recombinant SOD protein was used. The recombinant protein was secreted into the YPD culture medium as a glycosylated protein with a 32 mg/l expression yield. The purified recombinant protein possesses a specific activity of 440 U/mg.</p> <p>Conclusion</p> <p><it>D. antarctica </it>Cu/ZnSOD recombinant protein was expressed in a constitutive system, and purified in a single step by means of an affinity column. The recombinant SOD was secreted to the culture medium as a glycoprotein, corresponding to approximately 13% of the total secreted protein. The recombinant protein Cu/ZnSOD maintains 60% of its activity after incubation at 40°C for 30 minutes and it is stable (80% of activity) between -20°C and 20°C. The recombinant SOD described in this study can be used in various biotechnological applications.</p> |
url |
http://www.biomedcentral.com/1756-0500/2/207 |
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