小型猪CYP3A88基因克隆与其他动物的序列比较

Cloning of CYP3A88 in Experimental Miniature Pig and Sequence Comparision with Other Animals

目的克隆我国资源小型猪品系巴马香猪肝脏中的CYP3A88基因,并进行生物信息学分析。方法应用RACE(Rapid Amplification of cDNA Ends)技术对其全长进行扩增,测序,利用Internet和GenBank数据库对其序列进行生物信息学分析。结果首次克隆并鉴定了我国资源小型猪品系巴马香猪肝脏中CYP3A88(GenBank登录号:EF625347)的编码区,获得大小为1965bp的全长cDNA,编码区长为1512bp,编码503个氨基酸;比较核苷酸序列,与小型猪CYP3A39相似性高达94%,而与人等其它动物的CYP3A相似性则在86%以下;推导和分析氨基酸序列表明,与小型猪CYP3A其它成员(CYP3A39、CYP3A29、CYP3A22)进行对比,其相似性分别为92%,89%,80%,而将小型猪与人的CYP3A分别比对,小型猪CYP3A88与人CYP3A4相似性最高,为77%;对其二级结构预测,它可能含12个α螺旋,4个β折叠;经NCBI上的CDD程序分析可知,其39～491氨基酸区域为P4503A亚家族保守结构区域;经聚类分析,小型猪和狗的CYP3A与人有较近的进化关系;通过同源建模法对其在线建模,人CYP3A4晶体结构作为其模建模型,得到了其经典的三维结构。结论在猪CYP3A家族四个基因中,CYP3A88在序列和高级结构上均与人CYP3A4的最为相似。

Objective To explore the structure and effection of CYP3A88 in experimental miniature pig. Method we used RACE （Rapid Amplification of cDNA Ends） technique to clone the full length cDNA of the coding region（CDS） of CYP3A88 in experimental miniature pig liver. Results we got the full length cDNA of the coding region（CDS） of CYP3A88（GenBank： EF625347）, and its coding region was 1512 bp, which consisted of 503 amino acids. CYP3A88 shared 94% nucleotide sequence identities and 9：2% protein sequence identities with CYP3A39. However, compaired with CYP3A in other animals including human, its nucleotide sequence identities were not over 86 %, and its protein sequence identities was 92 %, 89 % and 80 % to the other pig CYP3A（CYP3A39,CYP3A29,CYP3A22）, but 77% to human CYP3A4. Predicting its secondary structure, it might have 12 ahelices and 4βsheets. By CDD procedure on NCBI, it had the conserved domain of Cytochrome P450 3A subfamily between39 and 491 amino acids. CYP3A of Pig and dog kept the closest to human in their phylogenetic tree. Besides, by homology modelling Oll line, considering the crystal structure of human CYP3A4 as its model, we got the classic 3D structure of CYP3A88. Conclusion Miniature pig CYP3A88 was the most similar to human CYP3A4 in pig CYP3A.