Bovine liver cytochrome c oxidase has been isolated and the subunit structure of this preparation compared with that of the bovine heart enzyme. Of the 10 nuclear-coded subunits, 3 were different in the 2 tissue forms, having different migrations in sodium dodecyl sulfate-polyacrylamide gel electrophoresis, different antigenicities to antibodies made against the heart subunits, and different N-terminal amino acid sequences. Subunit ASA of heart begins with the N-terminal sequence of SSG in liver and is different in 17 of the first 33 residues including a deletion of 2 residues in the liver isoform of this subunit. Subunit CVII of liver differs from its heart counterpart in 6 of the first 37 residues while subunit CIX from liver differs from the heart isoform in 15 of the first 25 residues. No differences between tissue types were observed in partial sequencing of the remaining nuclear-coded subunits. Recently, the major portion of the sequence of subunit CIX from rat liver has been obtained by cloning and sequencing of the cDNA for this polypeptide [Suske, G., Mengel, T., Cordingley, M., & Kadenbach, B. (1987) Eur. J. Biochem. 168, 233-237]. There is a greater sequence homology of the rat and bovine liver forms of CIX than there is between the bovine heart and liver isoforms.
There is a renewed interest in the structure and functioning of the mitochondrial respiratory chain with the realization that a number of genetic disorders result from defects in mitochondrial electron transfer. These socalled mitochondrial myopathies include diseases of muscle, heart, and brain. The respiratory chain can be fractionated into four large multipeptide complexes, an NADH ubiquinone reductase (complex I), succinate ubiquinone reductase (complex II), ubiquinol oxidoreductase (complex III), and cytochrome c oxidase (complex IV). Mitochondrial myopathies involving each of these complexes have been described. This review summarizes compositional and structural data on the respiratory chain proteins and describes the arrangement of these complexes in the mitochondrial inner membrane. This biochemical information is provided as a framework for the diagnosis and molecular characterization of mitochondrial diseases.
Skin fibroblasts were selected as having cytochrome-c oxidase deficiency by activity mesurements in whole cells. Each cell line was cultured in sufficient amount to isolate mitochondria for biochemical characterization. Cytochrome-c oxidase was then examined by activity measurements, by heme determination and by polypeptide analysis using antibodies specific to the enzyme subunits. The cytochrome-c ox&se activity in the different cell lines ranged from 9% to 54% of that of normal fibroblasts. Heme determinations and polypeptide analysis established that the lowered cytochrome-c oxidase activity was due to reduced amounts of the complex in the mitochondrial inner membrane. In all cases, there was defective assembly of the enzyme, with the amounts of mitochondrially coded and nuclear coded subunits being reduced proportionally. These studies show that fibroblasts can be used for prenatal diagnosis of mitochondrial diseases and are a useful system in which to study mitochondrial biogenesis.
A 25-year-old woman with Kearns-Sayre syndrome (KSS) had complete external ophthalmoplegia, short stature, ataxia, cardiac conduction defects, and pigmentary retinopathy. Muscle biopsy revealed ragged-red fibers. Electron microscopy showed increased numbers of mitochondria with disordered structure and paracrystalline inclusions. Enzymatic analysis revealed a deficiency of complex II of the mitochondrial respiratory chain, and, more specifically, a deficiency of succinic dehydrogenase, although both subunits of this enzyme proved to be present by immunologic analysis. Therapy with vitamin cofactors did not result in short-term improvement. This appears to be the first report of complex II deficiency in a patient with KSS.
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