摘要
甜菜碱高半胱氨酸甲基转移酶(BHMT,EC2.1.1.5)催化甜菜碱的甲基转移给高半胱氨酸(Hcy),而分别生成二甲基甘氨酸和蛋氨酸。利用RT-PCR和SMART RACE的方法从鲈鱼(Lateolabrax japonicus)肝脏中克隆了BHMT全长cDNA。该序列全长1461bp,5'端非翻译区72bp,3'端非翻译区183bp,开放阅读框1206bp,可编码一个由401个氨基酸组成的蛋白质,该蛋白质相对分子质量为44.32kD,等电点为7.21。氨基酸序列分析表明,BHMT具有较高的保守性,鲈鱼BHMT与人、小鼠等9个物种的同源性为77%～93%,其中与黄鲈(Percaflavescens)同源性最高,为93%。用RT-PCR分析BHMT基因在10个组织中的表达结果表明,只有在肝、肠和肾中有较高的表达。RT-PCR和定量PCR表明,鲈鱼从盐度25的海水转入盐度12的海水后,肝、肠和肾BHMT基因表达量有增加,而将鲈鱼从盐度为25的海水转入盐度为29的海水后,肝、肠和肾的BHMT基因表达则减少。腹腔注射甜菜碱可增加鲈鱼BHMT基因在肝、肠和肾三个组织中的相对表达量。这些结果表明,甜菜碱可诱导鲈鱼BHMT基因表达,而BHMT在适应鱼类环境渗透压变化中起重要作用。
Betaine homocysteine methyltransferase（BHMT,EC 2.1.1.5） catalyzes the transfer of a methyl group from betaine to homocysteines（Hcy） to form dimethylglycine and Met,respectively.A full-length cDNA of the BHMT in Lateolabrax japonicus was amplified using RT-PCR and SMART RACE methods.The cDNA of the BHMT in L.japonicus is 1 461 bp in size,with 72 bp 5＇-UTR,183 bp 3＇-UTR and 1206 bp ORF,encoding a protein of 401 amino acids with a molecular weight of 44.32 kD and pI 7.21.The sequence analysis indicated that the deduced amino acid sequence of BHMT shared high identity（77%-93%） with nine other species;the highest was 93% with Perca flavescens.Semi-quantitative RT-PCR was used to characterize the expression of BHMT in ten tissues including muscle,heart,eye,brain,gill,liver,intestine,kidney,adipose tissue and spleen.The results showed that BHMT is only expressed in the liver,intestines and kidney.BHMT mRNA in these three tissues declined after the fish were transferred from sea water to a higher salinity environment and induced when transferred to a lower salinity environment.BHMT gene in liver,intestine and kidney can also be induced after intraperitoneal injection of betaine.Our results show that betaine can induce the transcription of BHMT in fish,and BHMT play pivotal roles in adaptation to osmotic change.
基金
国家自然科学基金(30671608)
浙江省自然科学基金(M303345)
宁波市自然科学基金(2006A610088)
关键词
鲈鱼
BHMT
克隆
组织表达
盐度
Lateolabrax japonicus
BHMT
Clone
Tissues Expression
Salinity
