条斑紫菜谷胱甘肽S-转移酶基因的克隆与表达分析  

Molecular cloning and expression pattern of glutathione S-transferase gene in Porphyra yezoensis Ueda(Bangiales,Rhodophyta)

作　　者：周向红[1,2] 易乐飞[1,2] 李信书[1,2] 王萍[2] 阎斌伦[1] 

机构地区：[1]淮海工学院江苏省海洋生物技术重点实验室,江苏连云港222005 [2]淮海工学院海洋学院,江苏连云港222005

出　　处：《水产学报》2011年第9期1354-1361,共8页Journal of Fisheries of China

基　　金：江苏省海洋生物技术重点实验室开放课题(2008HS004);国家科技支撑计划重大项目(2006BAD09A01)

摘　　要：谷胱甘肽S-转移酶(glutathione S-transferase,GST)是一类多功能蛋白家族,主要参与解毒和抗氧化防御过程。为了研究GST在条斑紫菜叶状体解毒过程中的作用,克隆并分析了条斑紫菜一个可溶性谷胱甘肽S-转移酶基因(命名为PyGST)的基因组DNA序列和cDNA序列,采用实时荧光定量PCR研究了其在铅胁迫下的表达规律。PyGST包含一个长624 bp的完整开放阅读框,编码区内含有一个长248 bp的内含子。PyGST具有GST蛋白家族的保守碱基和保守结构域。PyGST与藻类GST的亲缘关系最近,与动物Sigma型GST的亲缘关系次之;在进化树上PyGST等大多数藻类GST与动物Sigma型GST聚为一簇,表明PyGST属类Sigma型GST。铅胁迫能显著诱导PyGST表达,说明在叶状体细胞内PyGST参与了重金属铅的解毒过程。Glutathione S-transferases（GSTs）are a superfamily of multifunctional proteins present in all organisms.GSTs catalyze the conjugation of reduced glutathioe with xenobiotics,to form more soluble,nontoxic compounds,ready to be excreted or compartmentalized.So GSTs are primarily involved in detoxification and antioxidant defense.So far,compared to higher plants and mammals,there is limited information on genes related to detoxification or antioxidant defense from Porphyra yezoensis Ueda.In order to investigate the contribution of GST to detoxification in thalli of P.yezoensis,molecular cloning and expression analysis of a glutathione S-transferase gene（designated as PyGST）from P.yezoensis were performed.The genomic DNA and cDNA sequences of PyGST were obtained with computer assisted cloning,and characterized using multiple bioinformatic programs.The relative mRNA expression levels of PyGST under lead stress were investigated in thalli of P.yezoensis using real-time quantitative PCR.The PyGST cDNA contained a 624 nt of continuous complete coding region,encoding a polypeptide of 207 amino acids with a calculated molecular mass of 22.6 ku.The alignment of the genomic DNA with the cDNA showed that PyGST contained a 248 bp of intron,whose ends were defined by the 5′-GT and 3′-AG rule.PyGST contained conserved N-and C-domains of GST family.The amino acids defining the binding sites of glutathione and xenobiotic substrates were also conserved.Sequence comparison of PyGST revealed 28.0%-39.6% and 23.0%-28.5% identity with most algal GSTs and animal Sigma class GSTs,respectively.PyGST was closely clustered with most algal GSTs in the phylogenetic tree.Phylogenetic tree also showed that most algal GSTs including PyGST were distinct from previously described GST classes,but were most closely related to the Sigma class.These features indicated that the PyGST belongs to Sigma-like GST.Lead stress significantly induced the expression of PyGST.The expression levels of PyGST induced by low concentrations of lead were hig

关 键 词：条斑紫菜 谷胱甘肽S-转移酶 克隆 表达 铅 

分 类 号：S917[农业科学—水产科学] Q781[生物学—分子生物学]