Identification of an Intramitochondrially Synthesized Proteolipid Associated with the Mitochondrial ATPase Complex as the Product of a Mitochondrial Gene Determining Oligomycin Resistance in Aspergillus nidulans

An extranuclear oligomycin‐resistant mutant of Aspergillus nidulans was isolated and biochemically analyzed. The mutant grows slowly in the absence of oligomycin and contains an excess of cytochrome c . The ATPase complex solubilized from mutant mitochondria with Triton X‐100 is 80 times more resistant to oligomycin than the enzyme of the parental strain. The enzyme complexes of both strains were further purified to electrophoretic homogeneity and resolved by dodecylsulfate gel electrophoresis in the presence of 8 M urea into 16 polypeptide bands of apparent molecular weights between 65000 and 6000. Four proteins of apparent molecular weights 43000, 21000, 11000 and 6000 are synthesized on cycloheximide‐resistant mitochondrial ribosomes. The smallest subunit is selectively extracted from the purified enzyme by neutral chloroform/methanol (2/1). A proteolipid of identical electrophoretic mobility and site of synthesis is the major component of proteins extractable with the same solvent from whole mitochondrial membranes. This protein has been isolated from the mutant and its parental strain, and purified to apparent homogeneity. The proteins from both strains start with N ‐formyl‐methionine and end with valine. However, a second internal methionine residue present in the wild‐type protein is absent, together with some other amino acids, from the mutant protein. It is concluded that this small lipophilic protein is coded by a mitochondrial gene determining oligomycin resistance, is synthesized on mitochondrial ribosomes and is associated with the membrane sector of the ATPase complex. The mutational alteration of the protein not only confers oligomycin resistance to the ATPase complex but also causes a structural alteration of the mitochondrial inner membrane.