同位素标记相对和绝对定量技术筛选石房蛤毒素毒性分子学研究

Screening for saxitoxin proteins using iTRAQ

目的应用同位素标记相对和绝对定量(isobaric tags for relative and absolute quantitation,iTRAQ)技术筛选麻痹性贝类毒素(paralytic shellfish poisoning,PSP)中石房蛤毒素(saxitoxins,STX)的差异表达蛋白。方法以小鼠神经母细胞瘤N2a为受试对象,使用浓度分别为10-8 mol/L和10-9 mol/L的STX标准品和阳性对照NSP(10-10 mol/L)进行染毒。提取细胞蛋白、定量和酶解。iTRAQ标记后(其中114标记空白组、115标记浓度组1(STX10-8 mol/L)、116标记浓度组2(STX10-9 mol/L)、117标记阳性对照组(NSP10-10 mol/L)),进行LC-MS/MS分析。使用生物信息学方法对差异蛋白进行分析。结果与空白组相比,共筛选出184个差异表达蛋白。与PSP相关的蛋白共有22个。筛选出的差异表达蛋白参与10种生物学过程、4种分子类型。结论通过iTRAQ技术分析得到STX的差异表达蛋白,发现ABCB6、TLR8、TES、TIAM2可能是与STX毒素相关的蛋白分子。

Objective To screen the differentially expressed proteins of saxitoxins（STX）in paralytic shellfish toxins（PSP）by using isobaric tags for relative and absolute quantitation techniques（iTRAQ）. MethodsMurine neuroblastoma N2 acells were exposed to 10-8 mol/L and 10-9 mol/L STX standards as well as to 10-10 mol/L NSP positive control respectively.Cellular proteins were extracted,quantified and hydrolysed.2 DLCMS/MS analysis was performed after labeling with iTRAQ（114 for the blank control,115 for 10-8 mol/L STX group,116 for10-9 mol/L STX group,117 for the positive control）.Differential proteins were analyzed by bioinformatics methods. Results Compared with the control group,184 differentially expressed proteins were identified.A total of 22 proteins were associated with PSP.These differentially expressed proteins were involved in 10 biological processes and 4 molecular types. Conclusions Some of the differentially expressed proteins of STX identified by iTRAQ analysis as ABCB6,TLR8,TES and TIAM2 might be STX toxin related proteins.