The ND2 subunit is labeled by a photoaffinity analogue of asimicin, a potent complex I inhibitor

NADH: ubiquinone oxidoreductase (complex I) is the entry enzyme of mitochondrial oxidative phosphorylation. To obtain the structural information on inhibitor/quinone binding sites, we synthesized [H-3] benzophenone-asimicin ([H-3]BPA), a photoaffinity analogue of asimicin, which belongs to the acetogenin family known as the most potent complex I inhibitor. We found that [H-3] BPA was photo-crosslinked to ND2, ND1 and ND5 subunits, by the three dimensional separation (bluenative/doubled SDS-PAGE) of [H-3] BPA-treated bovine heart submitochondrial particles. The crosslinking was blocked by rotenone. This is the first finding that ND2 was photo-crosslinked with a potent complex I inhibitor, suggesting its involvement in the inhibitor/quinone-binding. (C) 2010 Federation of European Biochemical Societies. Published by Elsevier B. V. All rights reserved.