野生型与栽培型太子参的iTRAQ定量蛋白质组学研究

iTRAQ-based quantitative proteomics of cultivated Pseudostellaria heterophylla and its wild type

采用同位素标记相对和绝对定量(i TRAQ)技术对野生型与栽培型太子参进行定量蛋白质组学研究,探讨两种生态型太子参蛋白质组表达水平的差异。所提取的太子参蛋白质样品经FASP酶解、i TRAQ试剂标记、高p H-RPLC分离、RPLC-MS分离分析,获取的串联质谱数据通过Protein Pilot 5.0软件搜库进行蛋白质鉴定,通过蛋白质相对定量的比较寻找差异表达蛋白;再对差异蛋白质进行GO(gene ontology)、KEGG代谢通路和STRING网络通路分析。共鉴定到3 775个蛋白质,其中3 676个蛋白质具有定量信息,与野生型相比,栽培型太子参上调差异蛋白质127个,下调差异蛋白质205个;生物信息学分析得到71个目标差异蛋白,主要分成9类:热休克蛋白、转移酶、氧化还原酶、裂解酶、异构酶、连接酶、水解酶、微管蛋白和移位酶。结果显示:野生型太子参的碳水化合物和氨基酸代谢功能较栽培型太子参强,而抗应激能力较栽培型太子参弱;GWD1、PHS1、GBE1、PGM和BAM1可能是调节不同生态型太子参中蔗糖变化的5种关键蛋白,met E和CYS可能是导致不同生态型太子参氨基酸差异的2种关键蛋白。本研究为解析不同生态型太子参次生代谢物差异的成因及其药材品质形成的蛋白质机制提供基础资料。

This work was designed to investigate proteins differentially expressed in cultivated Pseudostellaria heterophylla and its wild type using i TRAQ proteomics approach. The extracted proteins were digested using FASP method and identified by i TRAQ coupled with LC-MS/MS technology and then analyzed by Protein Pilot 5.0 search engine. Proteins differentially expressed were searched through comparison of relatively quantified proteins. The analysis was conducted using GO（gene ontology）, KEGG and STRING. A total of 3 775 proteins were detected, among them, 3 676 proteins can be quantified, of which 127 proteins were up-regulated and 205 were down-regulated in cultivated Pseudostellaria heterophylla. We found 71 significantly differentially expressed proteins for further analysis. These proteins were classified into nine categories： heat shock proteins, transferases, oxidoreductases, lyases, isomerases, ligases, hydrolases, tubulin and translocases. The results indicated that the carbohydrate and cellular amino acids metabolism of cultivated Pseudostellaria heterophylla were weaker than its wild type and its ability of responding to stress was much stronger. GWD1, PHS1, GBE1, PGM, and BAM1 are the important proteins to regulate sucrose; met E and CYS are the key proteins that regulate amino acids in cultivated and wild Pseudostellaria heterophylla. This will provide the basic information for exploring the cause of secondary metabolites differences in different ecotype of Pseudostellaria heterophylla and the protein mechanism of its quality formation.