title: Cloning, overexpression, purification, and characterization of a new iron superoxide dismutase from Jatropha curcas.
abstract: A novel iron superoxide dismutase (Fe-SOD) gene from Jatropha curcas was cloned and expressed in Escherichia coli BL21 (DE3). This recombinant enzyme was isolated by a combined procedure involving immobilized metal-ion affinity chromatography and ion-exchange chromatography. The apparent molecular mass of this purified enzyme (designated as JcFe-SOD) was 35 kDa on SDS-PAGE. The full-length complementary DNA sequence of JcFe-SOD contained a 918-bp open reading frame encoding a 305-amino-acid precursor of 34.589 kDa. The result of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry showed that the purified enzyme may own two forms: a dimer and a monomer. The enzyme was relatively stable and showed 54% activity when incubated in 70°C for 60 Min. JcFe-SOD was found to have good pH stability in the pH range of 5.5-9.5 at 25°C over 1 H incubation. The activity of this enzyme was gradually inhibited by increasing concentration of H₂O₂, 2-mercaptoethanol, and ethylenediaminetetraacetic acid. An assay of the atomic absorption spectrum showed the presence of 0.41 atom of Fe in each SOD subunit.
authors.affiliation: College of Life Sciences, Sichuan University, Chengdu, People's Republic of China.
authors.forename: Chao
authors.initials: C
authors.lastname: Ou-Yang
authors.forename: Feng
authors.initials: F
authors.lastname: Cai
authors.forename: Shun
authors.initials: S
authors.lastname: Gao
authors.forename: Bei
authors.initials: B
authors.lastname: Niu
authors.forename: Shenghua
authors.initials: S
authors.lastname: Wang
authors.forename: Fang
authors.initials: F
authors.lastname: Chen
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D004795:Enzyme Stability; D015870:Gene Expression; D006863:Hydrogen-Ion Concentration; D031286:Jatropha / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D013482:Superoxide Dismutase / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification* / Q000378:metabolism; D013696:Temperature
chemical_list: D018076:DNA, Complementary; D011994:Recombinant Proteins; D013482:Superoxide Dismutase