title: Complete Amino Acid Sequence of a Copper/Zinc-Superoxide Dismutase from Ginger Rhizome.
abstract: Superoxide dismutase (SOD) is an antioxidant enzyme protecting cells from oxidative stress. Ginger (Zingiber officinale) is known for its antioxidant properties, however, there are no data on SODs from ginger rhizomes. In this study, we purified SOD from the rhizome of Z. officinale (Zo-SOD) and determined its complete amino acid sequence using N terminal sequencing, amino acid analysis, and de novo sequencing by tandem mass spectrometry. Zo-SOD consists of 151 amino acids with two signature Cu/Zn-SOD motifs and has high similarity to other plant Cu/Zn-SODs. Multiple sequence alignment showed that Cu/Zn-binding residues and cysteines forming a disulfide bond, which are highly conserved in Cu/Zn-SODs, are also present in Zo-SOD. Phylogenetic analysis revealed that plant Cu/Zn-SODs clustered into distinct chloroplastic, cytoplasmic, and intermediate groups. Among them, only chloroplastic enzymes carried amino acid substitutions in the region functionally important for enzymatic activity, suggesting that chloroplastic SODs may have a function distinct from those of SODs localized in other subcellular compartments. The nucleotide sequence of the Zo-SOD coding region was obtained by reverse-translation, and the gene was synthesized, cloned, and expressed. The recombinant Zo-SOD demonstrated pH stability in the range of 5-10, which is similar to other reported Cu/Zn-SODs, and thermal stability in the range of 10-60 °C, which is higher than that for most plant Cu/Zn-SODs but lower compared to the enzyme from a Z. officinale relative Curcuma aromatica.
authors.affiliation: Department of Bioscience, School of Agriculture, Tokai University, 9-1-1 Toroku, Higashi-ku, Kumamoto, 862-8652, Japan.
authors.forename: Yuki
authors.initials: Y
authors.lastname: Nishiyama
authors.affiliation: Department of Advanced Bioscience, Kinki University, 3327-204 Nakamachi, Nara, 631-8505, Japan.
authors.forename: Tamo
authors.initials: T
authors.lastname: Fukamizo
authors.affiliation: Department of Bioscience, School of Agriculture, Tokai University, 9-1-1 Toroku, Higashi-ku, Kumamoto, 862-8652, Japan.
authors.forename: Kazunari
authors.initials: K
authors.lastname: Yoneda
authors.affiliation: Department of Bioscience, School of Agriculture, Tokai University, 9-1-1 Toroku, Higashi-ku, Kumamoto, 862-8652, Japan. araki@agri.u-tokai.ac.jp.
authors.forename: Tomohiro
authors.initials: T
authors.lastname: Araki
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D003300:Copper / Q000737:chemistry; D004795:Enzyme Stability; D020939:Zingiber officinale / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D027343:Rhizome / Q000201:enzymology*; D016415:Sequence Alignment; D013482:Superoxide Dismutase / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D053719:Tandem Mass Spectrometry; D013696:Temperature; D015032:Zinc / Q000737:chemistry
chemical_list: D011994:Recombinant Proteins; D003300:Copper; D013482:Superoxide Dismutase; D015032:Zinc
keywords: Amino acid sequencing; Cu/Zn-superoxide dismutase; Ginger rhizome; Zingiber officinale