大黄鱼(<i>Pseudosciaena crocea</i>)细胞色素P450 CYP4F7基因的克隆及表达分析

娄绘芳; 黄艳青; 吴信忠

大黄鱼(Pseudosciaena crocea)细胞色素P450 CYP4F7基因的克隆及表达分析

浙江大学动物科学学院, 浙江杭州, 310029

基金项目: 国家自然科学基金资助项目((30671619);浙江省重点科技计划项目(2004C23041)

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出版历程
- 收稿日期: 2007-10-11
- 修回日期: 2008-06-12

Molecular cloning, characterization of a homologue of cytochrome P450s 4F7 in Pseudosciaena crocea

College of Animal Sciences, Zhejiang University, Hangzhou 310029, China

摘要

摘要: CYP4F7是细胞色素P450超基因家族中CYP4F亚家族中的同工酶之一,而细胞色素P450在通过电子传递参与生物体代谢和转化内源和外源化合物方面起着重要的作用。以本实验室构建的大黄鱼消减杂交cDNA文库中623bp的CYP4F7片段设计引物,以大黄鱼的总RNA为模板,利用RACE-PCR的方法,克隆鉴定出了大黄鱼细胞色素P450 CYP4F7基因。结果表明:基因全长1934bp,5'端非编码区59bp,3'端非编码区264bp,开放阅读框1611bp,编码536个氨基酸。序列分析表明大黄鱼的CYP4F7氨基酸序列与舌齿鲈的相似性为91%。利用RT-PCR方法研究CYP4F7在大黄鱼各组织中的表达特征,结果表明CYP4F7在肝脏、脾脏中表达量最高,在心脏、肾脏、头肾、鳃丝中表达量次之。另外,CYP4F7在注射了细菌脂多糖的大黄鱼各组织中的表达量均低于正常的大黄鱼,认为细菌脂多糖可能是CYP4F7表达的抑制剂,说明CYP4F7可能与鱼类免疫反应有一定相关性。
- 大黄鱼 /
- 细胞色素P450 /
- CYP4F7 /
- 克隆 /
- 表达分析
Abstract: CYP4F7 is amember of the 4F subfamily of the Cytochrome P450,which plays a vital role in the course of either catalyzing and regulating the metabolism of endogenous substances or activating and degrading exogenetic substances by transferring electrons.A CYP4F7 cDNA sequence (GenBank accession number:EU 169141) was isolated and identified from the total RNA of Pseudosciaena crocea kidney using the combined methods of BD SMART^TM cDNA synthesis and RACE-PCR.The sequence of CYP4F7 cDNA is 1934 bp which contains a 5'UTR of 59bp and a 3'UTR of 264bp,and an open reading frame with 1611bp that encodes a protein of 536 amino acids.The amino acid sequence of P.crocea CYP4F7 shared 67%~71% of residue identity with the CYP4F subfamily of other high mammals such as human and mice,and shared the highest identity of 91% with sea bass,Dicentrarchuslabrax.Semi-quantitative RT-PCR analysis showed that mRNA expression of CYP4F7 can be detected in the spleen,heart,liver,anterior kidney,kidney and gills,but not in the intestine and muscle in the normal group and the lipopo lysaccharide (LPS)-stimulated group of P.crocea,in which the higher expression was found in the spleen and liver.The expression of the CYP4F7 in all these tissues examined showed some decrease in the LPS-treated P.crocea compared with normal P.crocea,suggesting that LPS might be an inhibit or of cytochrome P450 CYP4F7 gene and CYP4F7 might be related to large yellow croaker P.crocea immune response.
- Pseudosciaenacrocea /
- cytochrome P450 /
- CYP4F7 /
- molecular cloning /
- characterization

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参考文献(22)

冷欣夫,邱星辉.细胞色素P450酶系的结构、功能与应用前景[M].北京:科学出版社,2001.

HOCHACHKA P W,MOMMSEN T P,昌永华,等.鱼类的生物化学与分子生物学,第二卷-分子生物学前沿[M].北京:中国农业出版社,2003:113-130.

YASUSHI K,KUSUNOSE E,SUMINOTO H,et al.Purification and characterization of recombinant human neutrophil leukotfiene B4 v-Hydroxylase(Cytochrome P450 4F3)[J].Arch Biochem Biophys,1998,355(2):201-205.

JENA-HUGUES P,EDUARD N L,FALCK J R,et al.Evaluation of cytochrome P450 4 family as mediator of phospholipase Dectivation in aortic vascular smooth muscle cells[J].Life Scien,2005,77(9):1015-1029.

SABOURAULT C,AMICHOT M,PAURON D,et al.Cloning of CYP4F7,a Kidney-Specific P450 in the Sea Bass Dicentrarchus labrax[J].Biochem Biophys Res Commun,1999,258(1):155-161.

JOHAN BYLUND,NIKLAS F,ERNST H O.Gene expression of a novel cytochrome P450 of the CYP4F subfamily in human seminal vesicles[J].Biochem Biophys Res Commun,1999,261(1):169-174.

STURM A,HANSEN D.Altered cholinesterase and monooxygenase leves in Daphnia magna and Chironomus riparius esposed to environmental pollutants[J].Ecotoxicol Environ Safety,1999,42(1):9-15.

DONALD R.BULER J-L,WANG B.Rainbow trout cytochrome P450s:purification,molecular aspects,metabolic activity,induction and role in environmental monitoring1[J].Comp Biochem Phys:Part C,1998,121(3):107-137.

MUNTANE J,LONG V,MITJAVILA M T,et al.Effect of carrageenan-induced granuloma on hepatic cytochrome P-450 isozymes in rats[J].Inflammation,1995,19(2):143-156.

JOHN M T.Isolation and characterization of the cDNA encoding the channel catfish(Ictalurus punctatus)form of cytochrome P450[J].Gen Comp Endocrinol,1994,95(2):155-168.

AUINSDH K.ANSKK S,BROMMER C L,et al.Catalytic characterization and cytokine mediated regulation of cytochrome P450 4Fs in rat hepatoeytes[J].Archi Biochem biophy,2007,461(1):104-112.

CUI Xiao-ming,HENRY W S.Cloning and characterization of the rat cytochrome P450 4F5(CYP4FS)gene[J].Gene,2002,297(1-4):179-187.

张崇文,于涟,毛芝鹃,等.哈维氏弧菌外膜蛋白OmpK基因的克隆及原核表达[J].水产学报,2006,30(1):9-14.

鄢庆枇,苏永全,王军,等.网箱养殖大黄鱼弧菌病研究[J].集美大学学报(自然科学版),2001,6(3):1-6.

袁思平,王国良,金珊.海水网箱养殖大黄鱼诺卡氏菌病及防治[J].水产科学,2005,24(9):35-36.

GIBSON-KUEH S,NETTO P,et al.The Pathology of Systemic Iridoviral Disease in Fish[J].Journ Compar Pathol,2003,129(2-3):111-119.

ZHENG W B,LIU G Z,AO J Q,et al.Expression analysis of immune-related gene in the spleen of large yellow croaker(Pseudosciaena crocea)stimulated with poly I:C[J].Fish &Shellfish Immunol,2006,21(4):414-430.

HUANG Yan-qing,WU Xin-zhong.The analyses of expressed genes in head kidney of large yellow croaker,Pseudosciaena crocea stimulated with peptidoglycan by suppression subtractive hybridization[C]//第二届亚洲网箱水产养殖国际大会论文摘要集(CAA2).杭州:浙江大学,2006:85.

SAMBROOK J,FRITSCH E F,MANlATIS T.Molecular Cloning:A Laboratory Manual,3rd ed[M].北京:科学出版社,2002.

张成岗,贺福初.生物信息学方法与实践[M].北京:科学出版社,2002:63-93.

THOMPSON J D,HIGGINS D G,GIBSON T J.Clustal W:improving the sensitivity of progressive multiple sequence alignment through sequence weighting,position-specific gap penalties and weight matrix choice[J].Nuc Acids Res,1994,22:4673-4680.

CHEN J,MURRAY M,LIDDLE C,et al.Down-regulation of male-specific cytochrome P450s 2C11 and 3A2 in bile duct-ligated male rats:Importance to reduced hepatic content of cytochrome P450 in cholestasis[J].Hepatology,1995,22:580-587.

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