Resolution of mitochondrial and chloroplast membrane protein complexes from green leaves of potato on blue-native polyacrylamide gels

Purification of mitochondria and mitochondrial protein complexes from green tissues is often severely impaired by the presence of chloroplasts and the ir proteins. Here we present a method which allows analysis of respiratory protein complexes from potato leaves. The procedure includes the preparatio n of an organellar fraction specifically enriched in mitochondria and the s eparation of organellar protein complexes by blue-native polyacrylamide gel electrophoresis (BN-PAGE). For the first time mitochondrial and chloroplas t protein complexes have been resolved simultaneously in a native gel. BN-P AGE allowed the separation of eleven bands, including the mitochondrial NAD H-dehydrogenase, the bc(1) complex and the mitochondrial F-1-ATP synthase a s well as the chloroplast F-1-ATP synthase, the cytochrome b(6)f complex, t he two photosystems and the light harvesting complex. The resolution of the protein complexes in the first dimension was good enough to allow identifi cation of all subunits of individual complexes in the second dimension unde r denaturing conditions. Thus, BN-PAGE offers an opportunity to analyze mit ochondrial and chloroplast protein complexes from a single preparation from very small amounts of tissue. The implications of our findings, for studie s on protein expression and turnover in different tissues and developmental stages, are discussed.