为了丰富芍药 CHI 基因研究的基础数据，首先利用 RACE 技术对芍药 CHI 基因的 CDS 区进行克隆，并通过EMBOSS 软件分析序列的密码子偏好性，其次利用生物信息学软件和在线数据库对芍药 CHI 进行蛋白结构与功能预测。结果表明，克隆获得芍药 CHI 基因 cDNA 序列长度为898 bp（GenBank 序列号：JN119872），芍药 CHI 基因偏好使用以 A ／U 结尾的密码子，28种偏好密码子（RSCU ＞1），其中偏好性较强的有 UUG、UGU、CAU、AGA、AGG、CGG（RSCU≥2），在密码子的使用频率上，芍药 CHI 基因与酵母、大肠杆菌等模式生物基因组的差异均大于拟南芥基因组。芍药 CHI为非跨膜亲水的稳定蛋白，无信号肽，属于非分泌蛋白；主要分布在细胞质（45．0％）和微体（42．5％）中；无糖基化位点，14个磷酸化位点，其中第20位氨基酸处存在一个典型的磷酸激酶 PKC，含有1个异构酶 Chalcone 保守结构域；二级结构包括α螺旋（39％）、延伸链（20％）、β-转角结构（13％）和无规则卷曲（27％）；三级结构呈β-三明治折叠形成的倒置花束。发现了一个异构酶催化活性区域和一个重要的特异性蛋白质激酶 PKC 位点，为今后深入研究 CHI 基因功能提供有利的基础资料，此外，拟南芥真核表达更适合作为芍药 CHI 基因的外源表达系统。
Cloning,Codon Usage Bias and Protein Structure and Function Prediction of CHI Gene in Paeonia lactiflora
In order to enrich basic data in Paeonia lactiflora CHI gene,the CDS region of Paeonia lactiflora CHI gene was cloned by RACE technology,further codon usage was analyzed by EMBOSS software,then the struc-ture and function of CHI protein was predicted by bioinformatics software and online databases.Results indicated that,the 898 bp full-length cDNA of CHI of Paeonia lactiflora(GenBank No.:JN1 19872 )was obtained.Paeonia lactiflora CHI gene has certain preference of codon usage with the terminal base A or U.There were twenty-eight bi-ased codons(RSCU >1 ),whose biased strongly codons were UUG,UGU,CAU,AGA,AGG and CGG(RSCU≥2). In codon usage frequency,the differences between CHI gene and Saccharomyces,E.coli genomes were more than Arabidopsis genome.CHI was non-transmembrane and hydrophilic protein with no signal peptide,which located mostly in cytoplasm(45.0%)and microbody(42.5%)by the result of subcellular level prediction and does not be-long to the secretory protein.CHI protein had no glycosylation sites and fourteen phosphorylation sites,including a PKC kinase located in 20th amino acid.Additionally,it had one conservative domain named chalcone.The seconda-ry structure of CHI protein contained 39% α-helices,20% extended strand,13% β-turn and 27% random coil. Tertiary structure presented an inversion bouquet by beta sandwich folding.In this study,we found one conservative domain responsible for catalytic activity of isomerase and an specific protein kinases PKC site,which will provide advantageous basic data for further studying CHI gene function.Moreover,the eukaryotic Saccharomyces expression systems may be more suitable for the expression of CHI gene.
Paeonia lactifloraCHI geneCodon usage biasProtein structure and function
国家科技期刊平台
NSTL
万方数据
维普
