Abstract
Respiratory supercomplexes are found in mitochondria of eukaryotic cells and some bacteria. A hypothetical role of these supercomplexes is electron channeling, which in principle should increase the respiratory chain efficiency and ATP synthesis. In addition to the four classic respiratory complexes and the ATP synthase, U. maydis mitochondria contain three type II NADH dehydrogenases (NADH for reduced nicotinamide adenine dinucleotide) and the alternative oxidase. Changes in the composition of the respiratory supercomplexes due to energy requirements have been reported in certain organisms. In this study, we addressed the organization of the mitochondrial respiratory complexes in U. maydis under diverse energy conditions. Supercomplexes were obtained by solubilization of U. maydis mitochondria with digitonin and separated by blue native polyacrylamide gel electrophoresis (BN-PAGE). The molecular mass of supercomplexes and their probable stoichiometries were 1200 kDa (I1:IV1), 1400 kDa (I1:III2), 1600 kDa (I1:III2:IV1), and 1800 kDa (I1:III2:IV2). Concerning the ATP synthase, approximately half of the protein is present as a dimer and half as a monomer. The distribution of respiratory supercomplexes was the same in all growth conditions. We did not find evidence for the association of complex II and the alternative NADH dehydrogenases with other respiratory complexes.
Highlights
Ustilago maydis is an aerobic organism that fully depends on oxidative phosphorylation for the supply of ATP, pointing to mitochondria as a key organelle in the physiology of this basidiomycete and a potential target to study their structure–function relationships.In previous reports from our laboratory, we described the presence of the four classic respiratory complexes as well as complex V or ATP synthase in U. maydis mitochondria [1]
When mitochondria from mammals, plants, or fungi are solubilized by dodecyl-β-Dmaltoside (DDM), the respiratory complexes are observed as individual monomers, except complex III, which is a functional dimer [5,26,27]
Contrary to the stability of the respiratory complexes of bovine heart mitochondria solubilized by DDM, in U. maydis, the absence of complex III2 below the ATP synthase band was notorious
Summary
In previous reports from our laboratory, we described the presence of the four classic respiratory complexes (complex I or NADH:ubiquinone oxidoreductase; complex II or succinate:ubiquinone oxidoreductase; complex III or ubiquinol:cytochrome c oxidoreductase; complex IV or cytochrome c oxidase) as well as complex V or ATP synthase in U. maydis mitochondria [1]. It has been reported that respiratory complexes are assembled into supramolecular structures called supercomplexes [4]. When these structures contain complexes I, III, and IV, they are called respirasomes, because the full complement of enzymes should support the transfer of electrons from NADH to oxygen [5]. Supercomplexes with different stoichiometries are found in mitochondria from different sources [5,6,7,8]
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