Arnold Janine, Shapiguzov Alexey, Fucile Geoffrey, Rochaix Jean-David, Goldschmidt-Clermont Michel, Eichacker Lutz Andreas
Center for Organelle Research (CORE), University of Stavanger, Stavanger, Norway.
Methods Mol Biol. 2014;1072:667-76. doi: 10.1007/978-1-62703-631-3_46.
Gel electrophoresis has become one of the most important methods for the analysis of proteins and protein complexes in a molecular weight range of 1-10(7) kDa. The separation of membrane protein complexes remained challenging to standardize until the demonstration of Blue Native PAGE in 1991 [1] and Clear Native PAGE in 1994 [2]. We present a robust protocol for high-resolution separation of photosynthetic complexes from Arabidopsis thaliana using lithium dodecyl sulfate as anion in a modified Blue Native PAGE (LDS-PAGE). Here, non-covalently bound chlorophyll is used as a sensitive probe to characterize the assembly/biogenesis of the pigment-protein complexes essential for photosynthesis. The high fluorescence yield recorded from chlorophyll-binding protein complexes can also be used to establish the separation of native protein complexes as an electrophoretic standard.
