iTRAQ结合2D LC-MS/MS技术在草菇不同生长发育时期蛋白质组分析中的应用

Comparative analysis of proteomic profile at different development stages of Volvariella volvacea by iTRAQ-coupled 2D LC-MSMS

摘要:

【目的】旨在采用iTRAQ标记结合二维液相色谱串联质谱技术对草菇不同生长发育阶段的差异蛋白质组进行研究。【方法】首先将提取的草菇不同生长阶段蛋白样品进行SDS-PAGE分析, 其次将经二维液相色谱串联质谱技术获取的串联质谱数据通过MASCOT软件搜库, 之后对鉴定蛋白质数据进行了主成分分析(Principal component analysis, PCA)、层次聚类(Hierarchy clustering)分析、K-均值(K-means)聚类和Gene Ontology (GO)注释分析。【结果】试验结果显示, 共计获得2 335个不同肽段, 鉴定到1 039个蛋白质, 其中1 030个蛋白质具有定量信息。在子实体阶段中显著上调蛋白质64个, 下调蛋白质150个。生物信息学分析表明, iTRAQ标记技术结合二维液相色谱串联质谱可对不同生长发育时期的草菇蛋白样品进行有效地分离和鉴定。【结论】这一研究结果为深入研究草菇乃至其他大型担子菌子实体形成和发育的分子机制提供借鉴。

Abstract:

[Objective] This work aims to investigate differential expression proteins at different development stages of Volvariella volvacea using isobaric tags for relative and absolute quantification (iTRAQ)-coupled two-dimensional liquid chromatography tandem mass spectrometry (2D LC-MS/MS) proteomics approach. [Methods] Firstly, the proteins extracted from V. volvacea at different development stages were analyzed by SDS-PAGE. Secondly, the tandem mass spcetrometry data obtained from 2D LC-MS/MS were used to search using MASCOT search engine. After that, principal component analysis, hierarchical clustering and Gene Ontology were used to analyze the detected results. [Results] Results showed that 1 039 protein groups, included a total of 2 335 unique peptides, were identified. Among them, 1 030 protein groups were provided quantitative information. Contrast to mycelia, 64 up-regulated and 150 down-regulated significantly differential proteins were found in fruiting bodies at different development stages of V. volvacea. Bioinformatics analysis revealed that iTRAQ-coupled 2D LC-MS/MS was a unique method for isolating and identifying protein groups of V. volvacea at different development stages. [Conclusion] It is helpful to insight into the molecular mechanism of the fruiting body formation and development of V. volvacea and other macro-basidiomycetes in the future.