Molecular cloning, characterization and expression analysis of a catalase gene in <i>Paphia textile</i>

WU Xiangwei; LI Jiakai; TAN Jing; LIU Xiande

doi:10.1007/s13131-016-0829-6

织锦巴菲蛤(Paphia textile)过氧化氢酶基因cDNA全长克隆、序列同源分析与组织表达

doi: 10.1007/s13131-016-0829-6

1 福建省海洋渔业资源与生态环境重点实验室, 集美大学水产学院, 厦门 361021;
2 云南农业大学动物科学技术学院, 昆明 650201

基金项目: The National Natural Science Foundation of China under contract No. 31172397; the New Century Excellent Talents of Fujian Province University under contract No. JA14167; the Open Research Fund Program of Fujian Provincial Key Laboratory of Marine Fishery Resources and Eco-environment under contract No. Z814041.

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- 收稿日期: 2015-05-26
- 修回日期: 2015-12-03

Molecular cloning, characterization and expression analysis of a catalase gene in Paphia textile

1 Fujian Provincial Key Laboratory of Marine Fishery Resources and Eco-environment, Fisheries College, Jimei University, Xiamen 361021, China;
2 Animal Science and Technology College, Yunnan Agricultural University, Kunming 650201, China

摘要

摘要: 过氧化氢酶是机体内一种重要的抗氧化酶，它能够通过清除体内多余的过氧化氢来保护生物机体免受各种形式的氧化性损伤。本文采用RT-PCR和cDNA末端快速扩增技术（rapid amplification of cDNA ends,RACE）首次克隆了织锦巴菲蛤过氧化氢酶（PtCAT）基因的cDNA全长序列。PtCAT基因cDNA全长1921bp，包含50bp的5'非翻译区（untranslated region,UTR）、349bp的3'UTR和1542bp的开放阅读框（open reading frame,ORF）。ORF区共编码513个氨基酸，分子量约为58.4kD，理论等电点为8.2。序列比对表明PtCAT含有一个高度保守的催化位点序列（⁶¹FNRERIPERVVHAKGAG⁷⁷），一个近端亚铁血红素配体序列（³⁵²RLFSYSDP³⁵⁹），三个参与催化反应的氨基酸残基（H⁷²,N¹⁴⁵,and Y³⁵⁶），以及一个过氧化物酶体目标信号序列（⁵¹¹AQL⁵¹³）。PtCAT与其它CAT氨基酸序列一致性为88～66%，相似性为54～89%。PtCAT基因在织锦巴菲蛤6个组织中均表达，其中在消化腺中表达量最高。并且，在高温刺激织锦巴非蛤6h、2h、2h后PtCAT基因分别在消化腺、性腺和心脏中的表达量显著上调。相反，高温刺激织锦巴菲蛤后的整个监测过程中PtCAT基因在外套膜、鳃、闭壳肌中显著下调。本文的研究结果表明PtCAT属于过氧化氢酶基因家族中的一员，它可能参与织锦巴菲蛤对外界有害环境因素的应激反应中。
- 织锦巴菲蛤 /
- 过氧化氢酶(CAT) /
- 克隆 /
- 序列分析 /
- 表达分析 /
- 高温胁迫
Abstract: Catalase is an important antioxidant protein that can protect organisms against various forms of oxidative damage by eliminating hydrogen peroxide. In this study, the catalase cDNA of Paphia textile (PtCAT) was cloned using RTPCR and rapid amplification of cDNA ends (RACE). PtCAT is 1921 bp long and consists of a 5'-UTR of 50 bp, a 3'-UTR of 349 bp, and an ORF of 1 542 bp that encodes 513 amino acids with a molecular weight of 58.4 kD and an estimated isoelectric point of 8.2. Sequence alignment indicated that PtCAT contained a highly conserved catalytic signature motif (⁶¹FNRERIPERVVHAKGAG⁷⁷), a proximal heme-ligand signature sequence (³⁵²RLFSYSDP³⁵⁹), and three catalytic amino acid residues (H⁷², N¹⁴⁵, and Y³⁵⁶). PtCAT also contains two putative N-glycosylation sites (³⁴NKT³⁶ and ⁴³⁷⁴³⁹) and a peroxisome-targeting signal (⁵¹¹AQL⁵¹³). Furthermore, PtCAT shares 53%-88% identity and 29%-89% similarity with other catalase amino acid sequences. PtCAT mRNA was present in all tested organs, including the heart, digestive gland, adductor muscle, gonad, gill, and mantle, but its expression was highest in the digestive gland. High-temperature-induced stress produced two expression patterns of PtCAT mRNA:first, an initial up-regulation followed by a down-regulation in the heart, digestive gland, and gonad and, second, consistent down-regulation in all other organs. These results demonstrate that PtCAT is a typical member of the catalase family and might be involved in the responses to harmful environmental factors.
- Paphia textile /
- catalase (CAT) /
- cloning /
- sequence analysis /
- expression analysis /
- high temperature stress

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织锦巴菲蛤(Paphia textile)过氧化氢酶基因cDNA全长克隆、序列同源分析与组织表达

doi: 10.1007/s13131-016-0829-6

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Molecular cloning, characterization and expression analysis of a catalase gene in Paphia textile

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