枸杞胆碱合成关键酶基因PEAMT的克隆及生物信息学分析

利用RT-PCR和RACE结合的方法,获得了枸杞磷酸乙醇胺N-甲基转移酶PEAMT基因cDNA全序列(命名为LbPEAMT).LbPEAMT cDNA全长1 863 bp,开放读码框1 497 bp,编码一个由498个氨基酸构成的多肽,理论分子量为56.53 kDa,等电点pⅠ为5.50.通过多种序列比对,该基因编码的氨基酸序列与番茄PEAMT氨基酸的相似性为93％.二级结构预测LbPEAMT蛋白由44.98％的α-螺旋、17.67％的延伸链、6.63％的β-转角和30.72％的不规则卷曲组成.LbPEAMT是一个亲水蛋白.含有2个S-腺苷甲硫氨酸依赖催化活性结构域,分别位于N端65-164 aa和C端290-392aa.每个活性区都包含有4个部分基序,分别为Ⅰ、post Ⅰ、post Ⅱ和post Ⅲ.LbPEAMT的这2个活性区在蛋白、磷脂和小分子甲基转移酶中都是相对保守的.

Cloning and Bioinformatics Analysis of Choline Biosynthetic Gene PEAMT in Lycium barbarum

Betaine plays an imporant role in plant responses to various environmental stresses.Phosphoethanolamine N-methyltransferas (PEAMT) is the key regulatory enzyme in the biosynthesis pathway of the precursor choline.In this study,a full-lengh cDNA of PEAMT gene (named LbPEAMT) was firstly isolated and characterized from the leaves of L.barbarum. Full length LbPEAMT was 1 808 bp,containing a 1 497 bp ORF and encoding 498 amino acids.Comparative and bioinformatics analyses revealed that the amino acid sequence encoded by this gene (LbPEAMT) was highly homologous (93%) to PEAMTs from other plant species (i.e.,tomato).The predicted second structure of this LbPEAMT protein was composed of 44.98% of alpha-helix,17.67% of extended chain,6.63% of beta-turn and 30.70% of irregular curl.LbPEAMT was a hydrophilic protein containing two of the S-adenosylmethion-ine-dependent catalyzing domains,which were located in the N-terminal of 65-164 aa and C-terminal of 290-392 aa,respectively.Each catalyzing domain contained four motifs named as I,post I,post II,and post III.However,the two catalyzing domains of LbPEAMT were relatively conservative in the structure of protein methyltransferase,phospholipid transferase and methyltransferase of small molecules.