Characterization of a subcomplex of mitochondrial NADH:ubiquinone oxidoreductase (complex I) lacking the flavoprotein part of the N-module.
Mitochondrial NADH:ubiquinone oxidoreductase is the largest and most complicated proton pump of the respiratory chain. Here we report the preparation and characterization of a subcomplex of complex I selectively lacking the flavoprotein part of the N-module. Removing the 51-kDa and the 24-kDa subunit resulted in loss of catalytic activity. The redox centers of the subcomplex could be reduced neither by NADH nor NADPH demonstrating that physiological electron input into complex I occurred exclusively via the N-module and that the NADPH binding site in the 39-kDa subunit and further potential nucleotide binding sites are isolated from the electron transfer pathway within the enzyme. Taking advantage of the selective removal of two of the eight iron-sulfur clusters of complex I and providing additional evidence by redox titration and site-directed mutagenesis, we could for the first time unambiguously assign cluster N1 of fungal complex I to mammalian cluster N1b.
Zickermann
Volker
V
Johann Wolfgang Goethe-Universität, Fachbereich Medizin, Molekulare Bioenergetik, Centre of Excellence Frankfurt Macromolecular Complexes, D-60590 Frankfurt am Main, Germany.
Zwicker
Klaus
K
Tocilescu
Maja A
MA
Kerscher
Stefan
S
Brandt
Ulrich
U
D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D002384:Catalysis; D004578:Electron Spin Resonance Spectroscopy; D042967:Electron Transport Complex I / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D005486:Flavin Mononucleotide / Q000378:metabolism; D005420:Flavoproteins / Q000235:genetics; D016297:Mutagenesis, Site-Directed; D017384:Sequence Deletion; D015007:Yersinia / Q000201:enzymology*
D001426:Bacterial Proteins; D005420:Flavoproteins; D005486:Flavin Mononucleotide; D042967:Electron Transport Complex I