纳米铜对大鼠肝脏毒性相关蛋白过氧化氢酶的分离鉴定及生物信息学分析

Isolation,Identification and Bioinformatics Analysis of CAT Protein Related with Hepatotoxity by Copper Nanoparticles in Rats

【目的】分离和鉴定纳米铜对大鼠肝脏毒性相关蛋白过氧化氢酶(catalase,CAT),探讨CAT在毒性发挥中的作用,为揭示纳米铜对肝脏毒性机制提供依据。【方法】应用2-DE技术和PDQuest 8.0软件在大鼠肝脏蛋白组中筛选纳米铜对肝脏毒性差异蛋白,经质谱鉴定后进行生物信息学分析。【结果】筛选到下调的差异蛋白点6602和7702与肝毒性相关,鉴定均为CAT蛋白;其性质稳定,有一定亲水性,无信号肽,定位于细胞质,可能属于非分泌性蛋白,含有过氧化氢酶活性位点64FDRERIPERVVHAKGAG80和过氧化氢酶亚铁血红素配合基位点354RLFAYPDTH362等功能位点;无规则卷曲、α螺旋和延伸链是其主要的二级结构元件,并预测了其三级结构图;同源性分析表明,大鼠的CAT与其它8个物种有较高同源性,并构建了CAT蛋白的系统进化树。【结论】纳米铜通过下调大鼠肝脏中CAT蛋白表达,引起肝细胞氧化应激损伤,可能是其发挥毒性作用的途径之一。

【Objective】In order to investigate the hepatotoxic mechanisms of nanoparticles copper,catalase（CAT） was isolated and identified from liver,and analyzed by bioinformatics,which is related with hepatotoxity induced by copper nanoparticles in rats.【Method】 The differential expression proteins related with hepatotoxity of copper nanoparticles were screened by 2-DE and PDQuest 8.0 software and then analyzed by bioinformatics after identified by MALDI-TOF-TOF MS through comparative proteomics strategy.【Result】The 6602 and 7702 spots of differentially expressed proteins were found to associate with hepatotoxity.They were identified as CAT protein which was located in the cytoplasm.This hydrophilic protein had no signal peptide,and was non-secreted protein.It also contained catalase active sites 64FDRERIPERVVHAKGAG80 and catalase heme ligand sites 354RLFAYPDTH362.The random coils,α-helices and extended chains were its main secondary structural elements,and the three-dimensional structure was predicted.Homology analysis showed that CAT had a high homology between rat and the other eight species,and the phylogenetic tree of CAT was constructed.【Conclusion】Copper nanoparticles could regulate down the CAT protein expression so as to induce oxidative stress injury in liver cells,which may be a pathway of copper nanoparticles to exert the hepatotoxic effects in rats.