Cloning and constitutive expression of Deschampsia antarctica Cu/Zn superoxide dismutase in Pichia pastoris
Abstract Background Deschampsia antarctica 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 ge...
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ftdoajarticles:oai:doaj.org/article:874819a9f2734c7082466c9592e6a78a 2023-05-15T13:48:16+02:00 Cloning and constitutive expression of Deschampsia antarctica Cu/Zn superoxide dismutase in Pichia pastoris Gidekel Manuel Moraga Ana Bravo Ramírez León A Dinamarca Jorge Navarrete Alejandro Sánchez-Venegas Jaime R 2009-10-01T00:00:00Z https://doi.org/10.1186/1756-0500-2-207 https://doaj.org/article/874819a9f2734c7082466c9592e6a78a EN eng BMC http://www.biomedcentral.com/1756-0500/2/207 https://doaj.org/toc/1756-0500 doi:10.1186/1756-0500-2-207 1756-0500 https://doaj.org/article/874819a9f2734c7082466c9592e6a78a BMC Research Notes, Vol 2, Iss 1, p 207 (2009) Medicine R Biology (General) QH301-705.5 Science (General) Q1-390 article 2009 ftdoajarticles https://doi.org/10.1186/1756-0500-2-207 2022-12-31T04:56:20Z Abstract Background Deschampsia antarctica 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 D. antarctica Cu/ZnSOD gene into Pichia pastoris and to characterize the heterologous protein. Findings The Copper/Zinc superoxide dismutase (Cu/ZnSOD) gene, SOD gene, was isolated from a D. antarctica by cDNA library screening. This SOD gene was cloned in the expression vector pGAPZαA and successfully integrated into the genome of the yeast P. pastoris 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. Conclusion D. antarctica 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. Article in Journal/Newspaper Antarc* Antarctic Antarctica Directory of Open Access Journals: DOAJ Articles Antarctic The Antarctic BMC Research Notes 2 1 207 |
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Directory of Open Access Journals: DOAJ Articles |
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English |
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Medicine R Biology (General) QH301-705.5 Science (General) Q1-390 |
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Medicine R Biology (General) QH301-705.5 Science (General) Q1-390 Gidekel Manuel Moraga Ana Bravo Ramírez León A Dinamarca Jorge Navarrete Alejandro Sánchez-Venegas Jaime R Cloning and constitutive expression of Deschampsia antarctica Cu/Zn superoxide dismutase in Pichia pastoris |
topic_facet |
Medicine R Biology (General) QH301-705.5 Science (General) Q1-390 |
description |
Abstract Background Deschampsia antarctica 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 D. antarctica Cu/ZnSOD gene into Pichia pastoris and to characterize the heterologous protein. Findings The Copper/Zinc superoxide dismutase (Cu/ZnSOD) gene, SOD gene, was isolated from a D. antarctica by cDNA library screening. This SOD gene was cloned in the expression vector pGAPZαA and successfully integrated into the genome of the yeast P. pastoris 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. Conclusion D. antarctica 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. |
format |
Article in Journal/Newspaper |
author |
Gidekel Manuel Moraga Ana Bravo Ramírez León A Dinamarca Jorge Navarrete Alejandro Sánchez-Venegas Jaime R |
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 Deschampsia antarctica Cu/Zn superoxide dismutase in Pichia pastoris |
title_short |
Cloning and constitutive expression of Deschampsia antarctica Cu/Zn superoxide dismutase in Pichia pastoris |
title_full |
Cloning and constitutive expression of Deschampsia antarctica Cu/Zn superoxide dismutase in Pichia pastoris |
title_fullStr |
Cloning and constitutive expression of Deschampsia antarctica Cu/Zn superoxide dismutase in Pichia pastoris |
title_full_unstemmed |
Cloning and constitutive expression of Deschampsia antarctica Cu/Zn superoxide dismutase in Pichia pastoris |
title_sort |
cloning and constitutive expression of deschampsia antarctica cu/zn superoxide dismutase in pichia pastoris |
publisher |
BMC |
publishDate |
2009 |
url |
https://doi.org/10.1186/1756-0500-2-207 https://doaj.org/article/874819a9f2734c7082466c9592e6a78a |
geographic |
Antarctic The Antarctic |
geographic_facet |
Antarctic The Antarctic |
genre |
Antarc* Antarctic Antarctica |
genre_facet |
Antarc* Antarctic Antarctica |
op_source |
BMC Research Notes, Vol 2, Iss 1, p 207 (2009) |
op_relation |
http://www.biomedcentral.com/1756-0500/2/207 https://doaj.org/toc/1756-0500 doi:10.1186/1756-0500-2-207 1756-0500 https://doaj.org/article/874819a9f2734c7082466c9592e6a78a |
op_doi |
https://doi.org/10.1186/1756-0500-2-207 |
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BMC Research Notes |
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2 |
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1 |
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207 |
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1766249052056322048 |