忽地笑加兰他敏生物合成代谢途径中COMT酶基因克隆及序列分析  

Cloning and sequence analysis of LaCOMT in the biosynthesis pathway of galanthamine in Lycoris aurea

作　　者：桂柳姿[1] 崔培梧[1] 潘清平 唐金凤[3] 鲁耀邦 

机构地区：[1]湖南中医药大学,湖南长沙410208 [2]湖南省中药现代化研究重点实验室,湖南长沙410208 [3]广东医学院 [4]中药药性与药效三级科研实验室,湖南长沙410208

出　　处：《湖南中医药大学学报》2013年第3期35-40,共6页Journal of Hunan University of Chinese Medicine

基　　金：湖南省自然科学基金重点项目(10JJ2024);湖南省教育厅科研基金资助项目(07A052);湖南省中药学重点学科资助;湖南中医药大学生物工程重点学科资助

摘　　要：目的克隆忽地笑加兰他敏合成代谢途径中COMT酶基因及序列分析。方法通过比较数据库中四种植物烟草、唐松草、玉米和罂粟的儿茶酚氧位甲基转移酶(COMT)基因的同源序列,设计简并引物,克隆出忽地笑COMT酶基因(LaCOMT)的部分序列,然后通过RACE技术获得LaCOMT的全长序列。结果该序列开放读码框长度为1101bp,编码366个氨基酸,且此氨基酸序列具有依赖S-腺苷-L-甲硫胺酸(SAM)的甲基转移酶蛋白的三个保守序列,并与高等植物COMT氨基酸序列(CAA52462.1、AAQ01670.1、AAD29844.1和NP_001106047.1)的同源性大于74%。LaCOMT蛋白的预测分子量为40 501.9,等电点pI为5.95。结论此研究所获得的碱基序列为COMT酶基因的全长序列,且其预测的三级结构与咖啡酸-3-氧位甲基转移酶单体三级结构的1kyzE模型相似,均为依赖S-腺苷-L-甲硫胺酸(SAM)的甲基转移酶。Objective To clone and analyze sequence of the catechol-O-methyhransferase （COMT） gene in Lyeoris aurea biosynthesis pathway. Methods A pair of degenerate primers were designed through comparing the sequences of catechol-O-methyltransferase （COMT） gene in four plants such as tobacco, meadow rue, corn and papaver, then the partial sequence of COMT gene was cloned by PCR and a full-length sequence of LaCOMT was obtained by RACE technique. Results The sequence length of open reading frame was 1101 bp, encoding 366 amino acids with three conserved sequence of S-adenosyl-L-methionine-dependent methyhransferases in plant. And the homology of the amino acid sequence of LaCOMT with other higher plants COMT （CAA52462.1, AAQ01670.1, AAD29844.] and NP_001106047.1） revealed more than 74%. The pre- dicted molecular weight of LaCOMT protein was 40501.9 and pI was 5.95. Conclusion Sequence of LaCOMT obtained in this study is a full-length one. And its predicted tertiary structure is simi- lar with lkyzE model of caffeic acid-3-O-methyltransferase enzyme monomer, both are S-adeno-syl-L-methionine-dependent methyhransferases.

关 键 词：忽地笑 加兰他敏 儿茶酚氧位甲基转移酶 基因克隆 生物信息学分析 

分 类 号：Q781[生物学—分子生物学]