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对线粒体蛋白质组的鉴定和分析有助于理解线粒体的功能和相关疾病的发病机制,包括能量代谢、凋亡、自由基产生、产热作用、钙离子信号通路等.本实验旨在鉴定人类肝脏线粒体蛋白质组中的抗原优势蛋白.用线粒体蛋白质作为免疫原,经过细胞融合、筛选和克隆,制备了240多个单克隆抗体杂交瘤细胞系.单克隆抗体识别的线粒体蛋白抗原通过人类肝脏cDNA表达文库筛选方法鉴定,相应的线粒体蛋白质的亚细胞定位通过免疫组化证实.发现了肝脏线粒体中6个抗原优势蛋白,分别被至少两种特异性的单克隆抗体所识别.这6个蛋白分别是乙酰辅酶A酰基转移酶(线粒体3-酮酯酰辅酶A硫解酶)2、醛脱氢酶1家族A1、氨甲酰磷酸合成酶1、二氢硫辛酰胺S乙酰转移酶(丙酮酸脱氢酶复合物的E2组分)、烯酰辅酶A水合酶1和羟基类固醇(11β)脱氢酶1.这些单克隆抗体有望应用于人类肝脏蛋白质组计划的相关研究,如去除优势蛋白、蛋白与蛋白之间相互作用的研究和验证等.
Identification and analysis of the mitochondrial proteome will help understand mitochondrial function and pathogenesis of related diseases including energy metabolism, apoptosis, free radical production, thermogenesis, calcium signaling, etc. This experiment aimed to identify human liver Over 240 monoclonal antibody hybridoma cell lines were prepared using mitochondrial protein as immunogen, cell fusion, screening and cloning. The mitochondrial protein antigen recognized by monoclonal antibody was expressed by human liver cDNA Library screening method identified the corresponding mitochondrial protein subcellular localization confirmed by immunohistochemistry found in liver mitochondria six antigen dominant protein, respectively, by at least two kinds of specific monoclonal antibodies identified.The six proteins were Acetyl-CoA acyltransferase (mitochondrial 3-ketoacyl-CoA thiolase) 2, aldehyde dehydrogenase 1 family Al, carbamyl phosphate synthetase 1, dihydrolipoamide S acetyltransferase E2 component of the hydrogenase complex), enoyl-CoA hydratase 1 and hydroxysteroid (11β) dehydrogenase 1. These monoclonal antibodies are expected to be applied to human liver Proteome research programs, such as the removal of the advantages of protein, protein-protein interaction between research and verification.