香樟3-羟基-3-甲基戊二酰辅酶A还原酶(CcHMGRs)基因家族的克隆及表达分析  

Cloning and expression analysis of 3-hydroxy-3-methylglutaryl coenzyme A reductase(CcHMGR) genes in Cinnamomum camphora(L.) Presl

作　　者：郑汉 虞慕瑶[2] 濮春娟 陈美兰 李福全[3] 申业 黄璐琦[2] ZHENG Han;YU Mu-yao;PU Chun-juan;CHEN Mei-Ian;LI Fu-quan;SHEN Ye;HUANG Lu-qi(.Tianjin University of Traditional Chinese Medicine,Tianjin 301617,China;State Key Laboratory of Dao-di Herbs,National Resource Center for Chinese Materia Medica,China Academy of Chinese Medical Sciences,Beijing 100700,China;Hulunbeier Mongolian Medical Hospital,Hulunbeier 021000,China)

机构地区：[1]天津中医药大学,天津301617 [2]中国中医科学院中药资源中心,道地药材国家重点实验室培育基地,北京100700 [3]呼伦贝尔蒙医医院,内蒙古呼伦贝尔021000

出　　处：《药学学报》2020年第1期152-159,共8页Acta Pharmaceutica Sinica

基　　金：国家自然科学基金资助项目(81573533)

摘　　要：3-羟基-3-甲基戊二酰辅酶A还原酶(3-hydroxy-3-methylglutaryl coenzyme A reductase, HMGR)是萜类甲羟戊酸途径(mevalonic acid pathway, MVA)上的第一个限速酶,是细胞质中萜类代谢途径中的重要调控位点。本研究以5种化学型的香樟(Cinnamomum camphora)作为实验材料,基于转录组数据,从香樟cDNA中克隆出2个HMGR基因,分别命名为CcHMGR1 (GenBank登录号:MN163055)和CcHMGR2 (GenBank登录号:MN163056)。基因包含开放阅读框(ORF)分别为1 689 bp和1 683 bp,编码562和560个氨基酸残基,生物信息学分析推测其分子质量为59.819 kDa和59.397 kDa,等电点(theoretical pI)为8.20和8.61,均不含信号肽,存在2个跨膜结构。结合蛋白质保守区以及进化树分析,证实该基因确为HMGR家族基因。利用实时荧光定量PCR检测, CcHMGR1和CcHMGR2在香樟5种化学型中的表达模式类似,均在油樟中的表达量要高于另外4种化学型;并且2个CcHMGRs基因均在根中表达量最高,枝中最低。本研究首次从香樟中克隆出了CcHMGRs cDNA全长,并揭示了CcHMGRs在5种化学型及不同组织中的差异表达,为香樟萜类化合物生物合成途径关键酶基因的挖掘提供研究基础。3-Hydroxy-3-methylglutaryl coenzyme A reductase(HMGR) is the first rate-limiting enzyme of terpenoid biosynthesis in the mevalonic acid pathway(MVA) pathway. It is an important regulatory site in terpenoids metabolism pathway in the cytoplasm. According to the transcriptome database of Cinnamomum camphora, two HMGRs named CcHMGR1(GenBank: MN163055) and CcHMGR2(GenBank: MN163056) were cloned by cDNA from C. camphora. The ORF of CcHMGR1 and CcHMGR2 is composed of 1 689 bp and 1 683 bp, respectively,encoding 562 and 560 amino acids. The bioinformatics analysis of CcHMGR1 and CcHMGR2 indicated that the molecular weight of the encoded protein is 59.819 kDa and 59.397 kDa, with a theoretically isoelectric point of8.20 and 8.61, respectively. There are 2 transmembrane structures without signal peptide existing in the encoded amino acid of CcHMGRs. The analysis of sequence alignment and phylogenetic tree showed that the CcHMGRs belonged to the HMGR family. The camphor is divided into five chemitypes, according to the main chemical compounds in C. camphora. The results of the real time PCR indicated that the expression level of CcHMGRs in Cineol type was higher than that in Linalool type, iso-nerolidol type, Camphor type and Borneol type. CcHMGRs expressed highest in roots and lowest in branches. In this study, the cDNA full length of CcHMGRs were cloned from C. camphora for the first time. Our results revealed that the expression level of CcHMGRs were different among five chemical types and different plant tissues, and the research provides foundation for further study of the terpenoids biosynthetic pathway in C. camphora.

关 键 词：香樟 3-羟基-3-甲基戊二酰辅酶A还原酶(CcHMGR) 生物信息学 化学型 表达分析 

分 类 号：R931[医药卫生—生药学]