Department of Biochemistry and Food Chemistry, Molecular Plant Biology, University of Turku, FI-20014 Turku, Finland.
Biochem J. 2011 Oct 15;439(2):207-14. doi: 10.1042/BJ20102155.
Gel-based analysis of thylakoid membrane protein complexes represents a valuable tool to monitor the dynamics of the photosynthetic machinery. Native-PAGE preserves the components and often also the conformation of the protein complexes, thus enabling the analysis of their subunit composition. Nevertheless, the literature and practical experimentation in the field sometimes raise confusion owing to a great variety of native-PAGE and thylakoid-solubilization systems. In the present paper, we describe optimized methods for separation of higher plant thylakoid membrane protein complexes by native-PAGE addressing particularly: (i) the use of detergent; (ii) the use of solubilization buffer; and (iii) the gel electrophoresis method. Special attention is paid to separation of high-molecular-mass thylakoid membrane super- and mega-complexes from Arabidopsis thaliana leaves. Several novel super- and mega-complexes including PS (photosystem) I, PSII and LHCs (light-harvesting complexes) in various combinations are reported.
基于凝胶的类囊体膜蛋白复合物分析是监测光合作用机器动态的一种有价值的工具。天然聚丙烯酰胺凝胶电泳（Native-PAGE）保留了复合物的成分，通常还保留了蛋白质复合物的构象，从而能够分析其亚基组成。然而，由于天然聚丙烯酰胺凝胶电泳和类囊体溶解系统的多样性，该领域的文献和实际实验有时会引起混淆。在本文中，我们描述了通过天然聚丙烯酰胺凝胶电泳分离高等植物类囊体膜蛋白复合物的优化方法，特别是：（i）去污剂的使用；（ii）溶解缓冲液的使用；和（iii）凝胶电泳方法。特别关注从小麦叶中分离高分子质量的类囊体膜超复合物和 mega 复合物。报道了几种新的超复合物和 mega 复合物，包括 PS（photosystem）I、PSII 和 LHCs（light-harvesting complexes）的各种组合。
