Studies on superoxide dismutase and oxidative stress response in fishes

鱼类超氧化物歧化酶与氧化应激反应的研究

• 批准号: 12660186
• 负责人: OSATOMI Kiyoshi
• 金额: $ 2.18万
• 依托单位: Nagasaki University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12660186/
• 关键词: Superoxide dismutase; SOD; Fish; Purification; Cloning; Fish disease; Bacterial infection; Oxidative stress; スーパーオキシドジムターゼ

Cu, Zn-superoxide dismutase (Cu, Zn-SOD) activities were compared in Japanese flounder Paralichthys olivaceus for four different kinds of bacterial infections. A significant increase in Cu, Zn-SOD levels was recorded in hepato-pancreas at the initial stages of Edwardsiella tarda (E. tarda) infection. It was suggested that the remarkable change of Cu, Zn-SOD activities was a characteristic phenomenon of E. tarda.Cu, Zn-SOD and Mn-SOD were purified to homogeneity from Japanese flounder hepato-pancreas. On SDS-PAGE, the purified Cu, Zn-SOD and Mn-SOD gave single protein bands with molecular mass of 17.8 kDa and 26 kDa under reducing conditions, respectively. The N-terminal ammo acid sequences were determined for 25 amino acid residues (Cu, Zn-SOD) and 19 residues (Mn-SOD), the sequences were compared with other SODs. Reverse transcription-polymerase chain reaction of RNA from Japanese flounder hepato-pancreas was performed using primers designed according to the amino acid sequences determined, and we determined primary structure of Japanese flounder Cu, Zn-SOD and Mn-SOD. Furthermore, recombinant Cu, Zn-SOD (rCu, Zn-SOD) was expressed in Escherichia coli and was recovered as an active form.

比较了4种细菌感染对牙鲆体内Cu，Zn-SOD活性的影响。在迟缓爱德华氏菌（Edwardsiellatarda，E.迟发性）感染。表明Cu，Zn-SOD活性的显着变化是大肠杆菌的特征现象。从牙鲆肝胰脏中分离纯化了Cu、Zn-SOD和Mn-SOD。SDS-聚丙烯酰胺凝胶电泳显示，Cu，Zn-SOD和Mn-SOD在还原条件下均为单一蛋白条带，分子量分别为17.8kDa和26 kDa。测定了25个氨基酸残基（Cu，Zn-SOD）和19个氨基酸残基（Mn-SOD）的N端氨基酸序列，并与其它SOD进行了比较。根据测定的氨基酸序列设计引物，对比目鱼肝胰RNA进行逆转录-聚合酶链反应，确定比目鱼Cu、Zn-SOD和Mn-SOD的一级结构。此外，重组Cu，Zn-SOD（rCu，Zn-SOD）在大肠杆菌中表达，并以活性形式回收。

项目成果

Osatomi, Kiyoshi: "Purification, N-terminal amino acid sequence, and some properties of Cu, Zn-superoxide dismutase from Japanese flounder (Paralichthys olivaceus) hepato-pancreas"Comparative Biochemistry and Physiology. Part B, 128. 751-760 (2001)

Osatomi、Kiyoshi：“来自日本牙鲆（Paralichthys olivaceus）肝胰腺的 Cu、Zn-超氧化物歧化酶的纯化、N 末端氨基酸序列和一些特性”比较生物化学和生理学。

K.Osatomi: "Purification and sequence of manganese superoxide dismutase from Japanese flounder (Paralichthys olivaceus) hepato-pancreas"Fisheries Science. 68. 1621-1622 (2002)

K.Osatomi：“来自日本牙鲆（Paralichthys olivaceus）肝胰腺的锰超氧化物歧化酶的纯化和序列”渔业科学。

Osatomi, Kiyoshi: "Changes in Cu, Zn-SOD activity in Japanese flounder Paralichthys olivaceus with bacterial infection"Nippon Suisan Gakkaishi. 68(2). 207-213 (2002)

Osatomi、Kiyoshi：“日本牙鲆牙鲆细菌感染时 Cu、Zn-SOD 活性的变化”Nippon Suisan Gakkaishi。

K.Osatomi,Y.Masuda,K.Hara,T.Ishihara: "Purification, N-terminal amino acid sequence, and some properties of Cu,Zn-superoxide dismutase from Japanese flounder (Paralichthys olivaceus) hepato-pancreas"Comparative Biochemistry and Physiology Part B. (印刷中). (

K.Osatomi、Y.Masuda、K.Hara、T.Ishihara：“来自日本牙鲆（Paralichthys olivaceus）肝胰腺的 Cu,Zn-超氧化物歧化酶的纯化、N 末端氨基酸序列和一些特性”比较生物化学和生理学 B 部分（正在出版）。

K.Osatomi: "Purification, N-terminal amino acid sequence, and some properties of Cu,Zn-super oxide dismutase from Japanese flounder (Paralichthys olivaceus) hepato-pancreas"Comparative Biochemistry and Physiology Part B. 128. 751-760 (2001)

K.Osatomi：“来自日本牙鲆（Paralichthys olivaceus）肝胰腺的 Cu,Zn-超氧化物歧化酶的纯化、N-末端氨基酸序列和一些特性”比较生物化学和生理学部分 B. 128. 751-760 (2001)