三角帆蚌GPX基因cDNA全序列的克隆及其编码蛋白的结构分析

doi:10.3724/SP.J.1005.2010.00360

遗传 ›› 2010, Vol. 32 ›› Issue (4): 360-368.doi: 10.3724/SP.J.1005.2010.00360

三角帆蚌GPX基因cDNA全序列的克隆及其编码蛋白的结构分析

李西雷¹, 汪桂玲¹ , 李家乐^1,2, 袁一鸣¹

1. 上海海洋大学省部共建水产种质资源发掘与利用教育部重点实验室, 上海 201306;
2. 上海市高校水产养殖学E研究院, 上海 201306

收稿日期:2009-08-24 修回日期:2009-12-31 出版日期:2010-04-20 发布日期:2010-03-24
通讯作者: 汪桂玲;李家乐 E-mail:glwang@shou.edu.cn;jlli@shou.edu.cn
基金资助:
国家重点基础研究发展规划(973计划)前期研究专项(编号：2009CB126000), 国家自然科学基金项目(编号：30871923), 上海市科委地方院校能力建设项目(编号：08390510100)和上海市水产养殖重点学科建设项目 (编号：Y1101)资助

Full-length cDNA cloning and encoding protein structure analysis of GPX in Hyriopsis cumingii

LI Xi-Lei¹, WANG Gui-Ling¹, LI Jia-Le^1,2, YUAN Yi-Ming¹

1. Key Laboratory of Exploration and Utilization of Aquatic Genetic Resources, Shanghai Ocean University, Ministry of Education, Shanghai 201306, China; 2. Aquaculture Division, E-Institute of Shanghai Universities, Shanghai 201306, China

Received:2009-08-24 Revised:2009-12-31 Online:2010-04-20 Published:2010-03-24
Contact: WANG Gui-Ling;LI Jia-Le E-mail:glwang@shou.edu.cn;jlli@shou.edu.cn

摘要/Abstract

摘要： 根据本实验室构建的三角帆蚌cDNA文库中已标注的EST序列, 利用cDNA末端快速扩增法(RACE)克隆了三角帆蚌(Hyriopsis cumingii)谷胱甘肽过氧化物酶(Glutathione peroxidase, GPX)基因cDNA全序列。序列分析表明, 该基因cDNA序列全长1286 bp, 包括5′端非翻译区(Untranslated Region) 39 bp、3′端非翻译区659 bp和开放阅读框(Open reading frame, ORF)588bp, 共编码195个氨基酸, 分子量约为22.2 kDa, 理论等电点为8.44, 属于含硒类GPX。该氨基酸序列具有GPX所有亚型中均高度保守的3个环状结构, 对酶的三级结构起稳定作用。在线分析结果表明: GPX的氨基酸序列不存在明显的疏水区, 也不存在信号肽序列。氨基酸相似性对比结果显示, 三角帆蚌GPX氨基酸序列与脊椎动物GPX-2及GPX-1的序列相似度较高, 为73.1%-80.8%, 与其他型GPX相似度较小, 相似度低于60%。构建的系统进化树显示三角帆蚌GPX与其他几种鱼类GPX聚为一类, 与其他已发表的几种软体动物GPX相距较远, 推测本实验克隆的三角帆蚌GPX基因和已发表的软体动物不属于同一种GPX类型。

关键词: 三角帆蚌, GPX基因, RACE, 编码蛋白

Abstract: Based on the EST sequence of mantle cDNA library of Hyriopsis cumingii, the complete cDNA of Hyriopsis cumingii GPX gene was cloned by rapid amplification of cDNA ends (RACE) in this paper. The GPX full-length cDNA sequence was 1286 bp including a 39 bp 5′-untranslated region, a 659 bp 3′-untranslated region and an open reading frame (ORF) 588 bp in length, which encodes for 195 amino acids with the predicted molecular weight of 22.2 kDa and the isoelectric point of 8.44. Molecular structure analysis revealed that GPX was Se-GPX. The amino acid sequences have three highly conserved loop structures that have a stabilizing effect on the tertiary structure of the enzyme. The amino acid sequence did not contain obvious hydrophobic domain and signal peptide. Homologous analysis of amino acid sequences indicated that the GPX of H. cumingii shared a high similarity with GPX-2 and GPX-1 of vertebrates (73.1%-80.8%), and low similarity with other types (< 60%). Phylogenetic trees showed that GPX of Hyriopsis cumingii was clustered together with GPX of some fishes and far away from GPX of several other published Molluscas. These results indicated GPX gene of H. cumingii cloned in this study was different from the GPX genes from other Molluscas.

Key words: Hyriopsis cumingii, glutathione peroxidase, RACE, encoding protein

李西雷，汪桂玲，李家乐，袁一鸣. 三角帆蚌GPX基因cDNA全序列的克隆及其编码蛋白的结构分析[J]. 遗传, 2010, 32(4): 360-368.

LI Xi-Lei, HONG Gui-Ling, LI Jia-Le, YUAN Yi-Ming. Full-length cDNA cloning and encoding protein structure analysis of GPX in Hyriopsis cumingii[J]. HEREDITAS, 2010, 32(4): 360-368.

参考文献

[1] 王咏梅. 自由基与谷胱甘肽过氧化物酶. 解放军药学学报, 2005, 21(5): 369–371. [2] 任洪林, 柳增善, 王克坚. 鲍免疫相关基因和蛋白的研究进展. 遗传, 2009, 31(4): 348–358. [3] Ursini F, Maiorino M, Brigelius-Flohé L, Baumann KD, Roveri A, Schomburg D, Flohé L. Diversity of glutathione peroxidases. Methods Enzymol, 1995, 252: 38–53. [4] Arthur J R. The glutathione peroxidases. Cell Mol Life Sci, 2000, 57(13–14): 1825–1835. [5] Jo PG, Choi YK, Choi CY. Cloning and mRNA expression of antioxidant enzymes in the Pacific oyster, Crassostrea gigas in response to cadmium exposure. Comp Biochem Physiol C Toxicol Pharmacol, 2008, 147(4): 460–469. [6] 王琳, 梁旭方, 刘秀霞, 程炜轩, 郁颖. 微囊藻毒素和脂多糖对鲢鱼和草鱼肝脏去毒相关基因活体诱导表达的影响. 暨南大学学报(自然科学与医学版), 2009, 30(1): 111–116. [7] 白桂芹, 成军, 张树林. 谷胱甘肽过氧化物酶与肝炎病毒蛋白关系的研究进展. 胃肠病学和肝病学杂志, 2004, 13(1): 82–84. [8] 王凡, 赵元凤, 吕景才, 刘长发. 铜污染对扇贝内脏团抗氧化酶活性的影响. 水产科学, 2008, 27(12): 622–624. [9] Doyen P, Bigot A, Vasseur P, Rodius F. Molecular cloning and expression study of pi-class glutathione S-transferase (pi-GST) and selenium-dependent glutathione peroxi-dase(Se-GPx) transcripts in the freshwater bivalve Dreissena polymorpha. Comp Biochem Physiol C Toxicol Pharmacol, 2008, 147(1): 69–77. [10] Doyen P, Vasseur P, Rodius F. Identification, sequencing and expression of the selenium-dependent glutathione peroxidase transcript in the freshwater bivalve Unio tumidus exposed to Aroclor 1254. Comp Brioche Physiol C Toxicol Pharmacol, 2006, 144(2): 122–129. [11] Bai Z, Yin Y, Hu S, Wang G, Zhang X, Li J. Identification of genes involved in immune response, microsatellite and SNP markers from expressed sequence tags generated from hemocytes of freshwater pearl mussel (Hyriopsis cumingii). Marin Biotechnol, 2009, 11(4): 520–530. [12] 汪桂玲, 李家乐. 淡水育珠蚌外套膜提取总RNA的改良方法. 生物技术通报, 2008, (z1): 356–357. [13] 王佳, 张利平, 杨联, 费春红, 王磊, 谢超, 吴建平. 山羊、绵羊MT-IV分子特性研究. 遗传, 2008, 30(12): 1591–1596. [14] Lambert C, Leonard N, de Bolle X, Depiereux E. ESyPred3D: Prediction of proteins 3D structures. Bioin-formatics, 2002, 18(9): 1250–1256. [15] Aumann KD, Bedorf N, Brigelius-Flohé R, Schomburg D, Flohé L. Glutathione peroxidases revisited-simulation of the catalytic cycle by computer-assisted molecular mod-eling. Biomed Environ Sci, 1997, 10(2–3): 136–155. [16] Yeh SP, Liu KF, Chiu ST, Jian SJ, Cheng W, Liu CH. Identification and cloning of a selenium dependent glu-tathione peroxidase from giant freshwater prawn, Macro-brachium rosenbergii. Fish Shellfish Immunol, 2009, DOI:10,1016/j.fsi. [17] Bae YA, Cai GB, Kim SH, Zo YG, Kong Y. Modular evo-lution of glutathione peroxidase genes in association with different biochemical properties of their encoded proteins in invertebrate animals. BMC Evol Biol, 2009, DOI: 10.1186/1471-2148-9-72. [18] 马森. 谷胱甘肽过氧化物酶和谷胱甘肽转硫酶研究进展. 动物医学进展, 2008, 29(10): 53－56. [19] Chu FF, Doroshow JH, Esworthy RS. Expression, charac-terization, and tissue distribution of a new cellular sele-nium-dependent glutathione peroxidases, GSHPx-GI. J Biol Chem, 1993, 268(4): 2571–2576. [20] Vernet P, Rock E, Mazur A, Rayssiguier Y, Dufaure JP, Drevet JR. Selenium-independent epididymis-restricted glutathione peroxidase 5 protein(GPX5) can back up fail-ing Se-dependent GPXs in mice subjected to selenium de-ficiency. Mol Reprod Dev, 1999, 54(4): 362–370. [21] Toppo S, Vanin S, Bosello V, Tosatto SCE. Evolutionary and structural insights into the multifaceted glutathione peroxidase(Gpx) superfamily. Antioxidants Redox Signal, 2008, 10(9): 1501–1513. [22] Dear TN, Campbell K, Rabbitts TH. Molecular cloning of putative odorant-binding and odorant-metabolizing pro-teins. Biochemistry, 1991, 30(43): 10376–10382. [23] Kryukov GV, Castellano S, Novoselov SV, Lobanov AV, Zehtab O, Guigó R, Gladyshev VN. Ch

编辑推荐

Metrics

www.chinagene.cn
备案号：京ICP备09063187号-4
总访问:,今日访问:,当前在线:

三角帆蚌GPX基因cDNA全序列的克隆及其编码蛋白的结构分析

Full-length cDNA cloning and encoding protein structure analysis of GPX in Hyriopsis cumingii

PDF (PC)

可视化

被引次数

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 15

编辑推荐

Metrics

[1]	梁巍，刘新星，张博. 《遗传》2012年第9期封面说明[J]. 遗传, 2012, 34(9): 1152-0.
[2]	杨红波，梁巍，刘新星，朱作言，林硕，张博. 应用双荧光报告系统检测斑马鱼microRNA的动态表达[J]. 遗传, 2012, 34(9): 1181-1192.
[3]	李西雷，汪桂玲，李家乐. 三角帆蚌GPX基因结构特征及抗性相关SNP的筛选[J]. 遗传, 2012, 34(11): 1456-1464.
[4]	赵伯阳，汪代华，徐刚毅，赵文伯，郑程莉. 山羊CAST基因Ⅱ型转录本的克隆及在山羊不同组织中的表达[J]. 遗传, 2011, 33(4): 358-364.
[5]	李明，赵巧辉，陈其新，刘孟洲，石晓卫. 家兔BMP7基因的克隆及其生物信息学分析[J]. 遗传, 2008, 30(7): 885-892.
[6]	金萍，张玮玮，黄惠芳，马飞 . 日本七鳃鳗(Lampetra japonica)Lyb基因的克隆与分析[J]. 遗传, 2008, 30(12): 1597-1602.
[7]	刘岗，仲大莲，刘雪兰，余为一. 应用RACE法克隆鸽恒定链基因的研究[J]. 遗传, 2008, 30(1): 77-80.
[8]	娜仁花，旭日干. 绵羊fertilin β基因编码区的钓取与结构分析[J]. 遗传, 2007, 29(8): 951-951―956.
[9]	王春国，宋文芹. 花椰菜细胞质雄性不育基因特异PCR标记的筛选[J]. 遗传, 2005, 27(2): 236-240.
[10]	卜友泉，罗绪刚，刘彬，李素芬. 一步3’RACE快速构建鸡MnSOD全长cDNA克隆Rapid[J]. 遗传, 2004, 26(4): 519-521.
[11]	王少丽，盛承发，乔传令. cDNA末端快速扩增技术及其应用*[J]. 遗传, 2004, 26(3): 419-423.
[12]	王邦俊，王强，张志刚，张劲松，李学刚. 利用RACE技术扩增大豆抗病基因同源cDNA 5′末端序列[J]. 遗传, 2003, 25(4): 425-427.
[13]	刘浩，Anand Ghanekar，Matthew Chan，Ming Feng Liu，刘永锋，Gary Levy. 猪FGL2基因cDNA末端序列检测及结构分析[J]. 遗传, 2003, 25(1): 17-21.
[14]	杜占文，刘立仁，张俊武. 用RACE结合cDNA文库筛选的方法获取新的锌指蛋白基因[J]. 遗传, 2002, 24(3): 329-331.
[15]	邱为民，张思仲，武辉，张戈，肖翠英. 一种新的cDNA 末端快速扩增获取全长cDNA的方法[J]. 遗传, 2001, 23(5): 480-481.