NDLI: Cloning, Expression, and Purification of a Cu/Zn Superoxide Dismutase from Jatropha curcas

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Cloning, Expression, and Purification of a Cu/Zn Superoxide Dismutase from Jatropha curcas

Content Provider	Semantic Scholar
Author	Ou-Yang, Chao Gao, Shun Ji Cai, Feng Chung, Tsair-Wang Wang, Sheng-Hua Chen, Fang
Copyright Year	2013
Abstract	We report cDNA cloning, expression, purification, and characterization of a novel Cu/ Zn superoxide dismutase (SOD) from Jatropha curcas leaves. The full-length cDNA of this SOD contained a 496-bp open-reading frame (ORF) encoding 162 amino acid residues. The recombinant plasmid containing the SOD coding sequence was introduced into Escherichia coli, and the SOD was expressed as a fusion protein. The recombinant SOD was purified from a high-density fed-batch culture using a combination of immobilized metal ion affinity chromatography (IMAC) and Sephadex G25 desalting chromatography. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and matrix-assisted laser desorption/ ionization time-of-flight mass spectrometry (MALDI-TOF MS) analysis indicated that the recombinant SOD was a monomeric protein with a molecular mass of approximately 16.4 kDa. Isoelectric focusing showed that this SOD was a basic protein with pI values of 7.04, 7.33, 8.62, and 8.77. The activity of the SOD was stable at 70 °C for 10 min, and in a broad pH range from 4 to 9. The presence of urea (up to 8 M), guanidinium chloride (up to 6 M), and 2-mercaptoethanol (up to 8 mM) had little effect on the activity. The activity decreased gradually with increasing concentrations of imidazole, hydrogen peroxide, and ethylenediaminetetraacetic acid (EDTA). Atomic absorption spectrometry showed the presence of 0.239 copper and 0.258 zinc atoms, respectively, in the SOD polypeptide
Starting Page	60
Ending Page	69
Page Count	10
File Format	PDF HTM / HTML
DOI	10.1515/znc-2013-1-209
PubMed reference number	23659174
Journal	Medline
Volume Number	68
Alternate Webpage(s)	http://www.znaturforsch.com/s68c/s68c0060.pdf
Alternate Webpage(s)	http://www.znaturforsch.com/ac/v68c/68c0060.pdf
Journal	Zeitschrift fur Naturforschung. C, Journal of biosciences
Language	English
Access Restriction	Open
Content Type	Text
Resource Type	Article

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