3-DIMENSIONAL STRUCTURE OF NADH-DEHYDROGENASE FROM NEUROSPORA-CRASSA BY ELECTRON-MICROSCOPY AND CONICAL TILT RECONSTRUCTION

NADH-dehydrogenase (Complex I) is the first complex of the mitochondri al respiratory chain. It is an amphipatic molecule located in the inne r mitochondrial membrane and is composed of at least 35 unique subunit s encoded by both mitochondrial and nuclear DNA. The whole complex was isolated in detergent from the fungus Neurospora crassa. It is very s table in its isolated form and was analysed as such by electron micros copy. Its mass, determined by dark-field scanning electron microscopy was estimated as 1.12 MDa. The complex was imaged by transmission elec tron microscopy, by negative staining and by cryo-electron microscopy. A three-dimensional model, with a resolution estimated at 35 Angstrom , was calculated from images of negatively stained complexes by the ra ndom conical tilt reconstruction technique. This model confirms the ge neral L-shape of the molecule, with arms of equal length and corrobora tes the hypothesis of a subdivision of the whole complex into three fu nctional domains. Immuno-labelling of the 49 kDA subunit of the periph eral arm allowed its localization within the complex. This is a first step in the subunit mapping of Complex I and the understanding of its activity. (C) 1997 Academic Press Limited.