New Insights into the Respiratory Chain of Plant Mitochondria. Supercomplexes and a Unique Composition of Complex II1

Holger Eubel, L. Jansch, H-P. Braun

Research output: Contribution to journalArticlepeer-review

286 Citations (Scopus)

Abstract

A project to systematically investigate respiratory supercomplexes in plant mitochondria was initiated. Mitochondrialfractions from Arabidopsis, potato (Solanum tuberosum), bean (Phaseolus vulgaris), and barley (Hordeum vulgare) werecarefully treated with various concentrations of the nonionic detergents dodecylmaltoside, Triton X-100, or digitonin, andproteins were subsequently separated by (a) Blue-native polyacrylamide gel electrophoresis (PAGE), (b) two-dimensionalBlue-native/sodium dodecyl sulfate-PAGE, and (c) two-dimensional Blue-native/Blue-native PAGE. Three high molecularmass complexes of 1,100, 1,500, and 3,000 kD are visible on one-dimensional Blue native gels, which were identified byseparations on second gel dimensions and protein analyses by mass spectrometry. The 1,100-kD complex represents dimericATP synthase and is only stable under very low concentrations of detergents. In contrast, the 1,500-kD complex is stable atmedium and even high concentrations of detergents and includes the complexes I and III2. Depending on the investigatedorganism, 50% to 90% of complex I forms part of this supercomplex if solubilized with digitonin. The 3,000-kD complex,which also includes the complexes I and III, is of low abundance and most likely has a III4I2 structure. The complexes IV,II, and the alternative oxidase were not part of supercomplexes under all conditions applied. Digitonin proved to be the idealdetergent for supercomplex stabilization and also allows optimal visualization of the complexes II and IV on Blue-nativegels. Complex II unexpectedly was found to be composed of seven subunits, and complex IV is present in two differentforms on the Blue-native gels, the larger of which comprises additional subunits including a 32-kD protein resembling COXVIb from other organisms. We speculate that supercomplex formation between the complexes I and III limits access ofalternative oxidase to its substrate ubiquinol and possibly regulates alternative respiration. The data of this investigation areavailable at http://www.gartenbau.uni-hannover.de/genetik/braun/AMPP.
Original languageEnglish
Pages (from-to)274-286
JournalPlant Physiology
Volume133
Issue number1
DOIs
Publication statusPublished - 2003

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