鲈鱼CPTⅠ基因cDNA克隆及表达分析

Molecular cloning and tissue distribution of carnitine palmitoyltransferases Ⅰ gene from Lateolabrax japonicus

肉毒碱棕榈酰转移酶(carnitine palmitoyltransferases,CPT,EC.2.3.1.2)在脂肪酸的β-氧化中起重要作用。线粒体内膜外侧的脂酰CoA经CPTⅠ催化与肉毒碱结合形成脂酰肉毒碱而得以穿过线粒体内膜,进入线粒体。在线粒体内膜内侧的CPTⅡ催化下,脂酰肉毒碱上的脂酰基又转移到CoA上,重新形成脂酰CoA,成为β-氧化的底物。利用RT-PCR和SMART RACE的方法从鲈鱼(Lateolabrax japonicus)肝脏中克隆了CPT I全长cDNA。该序列全长3007 bp,5'非翻译区166 bp、3'非翻译区477 bp、开放阅读框2364 bp,编码一个由787个氨基酸组成的蛋白质,分子量为89.60 kDa,等电点为8.85。氨基酸序列分析表明,鲈鱼CPT I具有较高的保守性,与金头鲷(Sparus aurata)、虹鳟(Oncorhynchus mykiss)、斑马鱼(Danio rerio)、人(Homo sapiens)、小鼠(Mus musculus)等7个物种的同源性为93%～66%,其中与金头鲷同源性最高,为93%。用RT-PCR分析CPT I基因在10个组织中的表达,结果表明在肌、肾中有较高的表达,心、脑、鳃、肝、肠次之,眼、脂肪、脾表达最低。

Carnitine palmitoyltransferases （CPT, EC. 2. 3. 1.2 ） plays an important role in the β-oxidation of the fatty acid. Acyl CoA is catalyzed into acyl earnitine by canaitine palmitoyltransferases I （ CPT I） on the outer mitoehondrial membrane. After transferred into inner mitochondrial membrane, acyl carnitine is converted to carnitine by CTP II. The cDNA of the CPT I in Lateolabrax japonicus is 3007 bp, with 166 bp 5＇-UTR, 477 bp 3＇-UTR and 2364 bp ORF, encoding a protein of 787 amino acids with a molecular weight of 89. 60k and pI 8.85. The sequence alignment analysis indicated that the deduced amino acid sequence of CPT I shared high identity （93% - 66% ） with seven other species; the highest was 93% with that of Sparus aurata. Semi-quantitative RT-PCR was used to characterize the expression of CPT 1 in ten tissues including muscle, heart, eye, brain, gill, liver, intestine, kidney, adipose tissue and spleen. The results showed that CPT I is mainly expressed in the muscle and kidney.