Proteomics of a toxic dinoflagellate Alexandrium catenella DH01:Detection and identification of cell surface proteins using fluorescent labeling  

作　　者：LI Cheng WANG DaZhi DONG HongPo XIE ZhangXian HONG HuaSheng 

机构地区：[1]State Key Laboratory of Marine Environmental Science,College of the Environment and Ecology,Xiamen University,Xiamen 361005,China

出　　处：《Chinese Science Bulletin》2012年第25期3320-3327,共8页

基　　金：supported by the National Natural Science Foundation of China (40876059);the National Key Basic Research Program of China(2010CB428703);the Doctoral Fund of Ministry of Education of China(20070384014),the Excellent Group and the Program for New Century Excellent Talents in University to D.Z. Wang

摘　　要：Alexandrium catenella DH01 is a toxic dinoflagellate species that is able to not only produce paralytic shellfish toxins,but also cause harmful algal blooms along the coast of China.In this study,we presented a new protocol for specific labeling and detection of the cell surface proteins(CSPs) of A.catenella DH01 cells using CyDye difference gel electrophoresis(DIGE) fluor minimal dyes.CSPs were identified using two-dimensional gel electrophoresis(2-DE) and MALDI TOF-TOF mass spectrometry(MS).The results showed that the fluorescent cyanine dye Cy3 could specifically label the CSPs of A.catenella DH01,with minimal labeling of intracellular proteins.Among three protein extraction methods evaluated,the Trizol method was the most efficient to extract CSPs with respect to protein spot number and resolution.Forty-one CSPs were separated and identified from A.catenella DH01 by 2-DE,in which 14 were identified in the protein database using MALDI TOF-TOF MS analysis.This work represents the first attempt to investigate the CSPs of A.catenella using the CyDye DIGE fluor dyeing method that provides a potentially important tool for future comprehensive characterization of CSPs and elucidation of the physiological functions of CSPs in dinoflagellates.Alexandrium catenella DH01 is a toxic dinoflagellate species that is able to not only produce paralytic shellfish toxins, but also cause harmful algal blooms along the coast of China. In this study, we presented a new protocol for specific labeling and detection of the cell surface proteins （CSPs） of A. catenella DH01 cells using CyDye difference gel electrophoresis （DIGE） fluor minimal dyes. CSPs were identified using two-dimensional gel electrophoresis （2-DE） and MALDI TOF-TOF mass spectrometry （MS）. The results showed that the fluorescent cyanine dye Cy3 could specifically label the CSPs of A. catenella DH01, with minimal labeling of intracellular proteins. Among three protein extraction methods evaluated, the Trizol method was the most efficient to extract CSPs with respect to protein spot number and resolution. Forty-one CSPs were separated and identified from A. catenella DH01 by 2-DE, in which 14 were identified in the protein database using MALDI TOF-TOF MS analysis. This work represents the first attempt to investigate the CSPs of A. catenella using the CyDye DIGE fluor dyeing method that provides a potentially important tool for future comprehensive characterization of CSPs and elucidation of the physiological functions of CSPs in dinoflagellates.

关 键 词：蛋白质组学 荧光标记 亚历山大 表面蛋白 细胞内 鉴定 检测 二维凝胶电泳 

分 类 号：Q51[生物学—生物化学] Q78