Schröder Lucie, Eubel Holger, Braun Hans-Peter
Institute of Plant Genetics, Plant Proteomics Group, Leibniz University Hannover, Hannover, Germany.
Methods Mol Biol. 2022;2363:101-110. doi: 10.1007/978-1-0716-1653-6_9.
Most molecular functions depend on defined associations of proteins. Protein-protein interactions may be transient or long-lasting; they may lead to labile assemblies or more stable particles termed protein complexes. Studying protein-protein interactions is of prime importance for understanding molecular functions in cells. The complexome profiling approach allows to systematically analyze protein assemblies of cells or subcellular compartments. It combines separation of intact protein fractions by blue native (BN) polyacrylamide gel electrophoresis (PAGE) and protein identification as well as quantification by mass spectrometry. Complexome profiling has been successfully applied to characterize mitochondrial fractions of plants. In a typical experiment, more than 1000 mitochondrial proteins are identified and assigned to defined protein assemblies. It allows discovering so far unknown protein complexes, studying assembly pathways of protein complexes and even characterizing labile super- and megacomplexes in the >10 mega-Dalton range. We here present a complexome profiling protocol for the straightforward definition of the protein complex inventory of mitochondria or other subcellular compartments from plants.
大多数分子功能依赖于蛋白质的特定关联。蛋白质-蛋白质相互作用可能是短暂的或持久的；它们可能导致不稳定的组装体或更稳定的颗粒，即所谓的蛋白质复合物。研究蛋白质-蛋白质相互作用对于理解细胞中的分子功能至关重要。复合物组分析方法能够系统地分析细胞或亚细胞区室中的蛋白质组装体。它结合了通过蓝色天然（BN）聚丙烯酰胺凝胶电泳（PAGE）分离完整蛋白质组分以及通过质谱进行蛋白质鉴定和定量。复合物组分析已成功应用于表征植物的线粒体组分。在一个典型实验中，可鉴定出1000多种线粒体蛋白质，并将其分配到特定的蛋白质组装体中。它能够发现迄今未知的蛋白质复合物，研究蛋白质复合物的组装途径，甚至表征分子量大于10兆道尔顿范围内不稳定的超级和巨型复合物。我们在此介绍一种复合物组分析方案，用于直接确定植物线粒体或其他亚细胞区室的蛋白质复合物清单。
