Assembly of cytochrome-c oxidase in cultured human cells

The assembly of cytochrome-c oxidase was studied in human cells cultured in the presence of inhibitors of mitochondrial or cytosolic protein synthesis. Mitochondrial fractions were resolved using two-dimensional PAGE (blue native PAGE and tricine/SDS/PAGE) and subsequent western blots were developed with monoclonal antibodies against specific subunits of cytochrome-c oxidase. Proteins were also visualized using metabolic labeling followed by two-dimensional electrophoresis and fluorography. These techniques allowed identification of two assembly intermediates of cytochrome-c oxidase. Assembly of the 13 subunits of cytochrome-c oxidase starts with the association of subunit I with subunit IV. Then a larger subcomplex is formed, lacking only subunits VIa and either VIIa or VII

Subunit IV of human cytochrome c oxidase, polymorphism and a putative isoform

As part of our study of isoenzyme forms of human cytochrome c oxidase, we purified subunit IV from human heart and skeletal muscle with reversed-phase HPLC and determined the N-terminal amino acid sequences and the electrophoretic mobility. The N-terminus of human heart subunit IV proved to be ragged with 30% of the protein lacking the first three residues. Also a Tyr/Phe polymorphism was observed at residue 16. No differences in N-terminal sequence and electrophoretic mobility were observed between subunit IV of cytochrome c oxidase from human heart and skeletal muscle. Therefore, our results suggest that identical subunits IV are present in cytochrome c oxidase from human heart and skeletal muscle. A putative isoform of subunit IV with a blocked N-terminus was purified from human heart cytochrome c oxidase, which proved to have a different retention time on a reversed-phase column and also a slightly higher electrophoretic mobility on an SDS-polyacrylamide gel compared to the native subunit IV. We could not demonstrate the existence of isoforms of subunit IV in human skeletal muscl

Demonstration of two isoforms of subunit VIIa of cytochrome c oxidase from human skeletal muscle. Implications for mitochondrial myopathies

Two different isoforms of subunit VIIa have been found in cytochrome c oxidase isolated from human skeletal muscle. The first 22 residues of the N-terminal amino acid sequences showed 5 differences. Our results provide the first conclusive evidence for the existence of cytochrome c oxidase isoenzymes in man. Since the two cytochrome c oxidase isoforms were both present in skeletal muscle tissue, though not necessarily in the same cell type, this suggests that human cytochrome c oxidase isoforms are not strictly tissue-specific. These findings may have important implications for the elucidation of genetic diseases in man in which a deficiency of cytochrome c oxidase is restricted to certain tissue

Home blood pressure monitoring detects unrevealed hypertension in women with a history of preeclampsia: results of the BP-PRESELF study

OBJECTIVES: The risk of cardiovascular disease more than doubles after hypertensive disorders of pregnancy. As early onset chronic hypertension contributes to cardiovascular risk, implementation of screening strategies, using home blood pressure monitoring (HBPM), may help to improve long-term cardiovascular health.We evaluated whether HBPM among women with a history of preeclampsia/HELLP syndrome is feasible for early detection and management of hypertension. METHODS: The BP-PRESELF study is a multicenter randomized controlled trial. Participants were randomized to intervention group with HBPM for the duration of 1 year or the control group with 'usual care'. The primary outcome was feasibility of HBPM during 1 year of follow-up, defined as protocol adherence, protocol persistence and patient acceptance. Secondary outcomes were blood pressure levels and prevalence of hypertension. RESULTS: We recruited 198 women with a mean age of 45 years. Protocol adherence decreased during the first 6 months, after which it stabilized. Protocol persistence remained high throughout follow-up. During the study period, 33 women (34%) in the intervention group were diagnosed with hypertension versus only 10 women (11%) in the control group, P<0.001. At 1-year follow-up, mean systolic blood pressure (SD) was 120.4 (11.6) mmHg in the intervention group versus 126.1 (14.3) mmHg in the control group, P=0.003. Mean diastolic blood pressure (SD) values were 77.1 (8.0) mmHg versus 81.7 (9.4) mmHg, P<0.001, respectively. Adjusted systolic and diastolic differences (95% confidence interval) were -6.81 (-10.17, -3.45) and -4.93 (-7.26, -2.61) mm Hg, with 80% less hypertension at 1-year follow-up in the intervention group. CONCLUSIONS: HBPM appears to be feasible for follow-up of blood pressure in women after preeclampsia/HELLP syndrome, while it detected hypertension and blood pressure levels reduced in one-third of women in this group

Refsum disease is caused by mutations in the phytanoyl-CoA hydroxylase gene

Refsum disease is an autosomal-recessively inherited disorder characterized clinically by a tetrad of abnormalities: retinitis pigmentosa, peripheral neuropathy, cerebellar ataxia and elevated protein levels in the cerebrospinal fluid (CSF) without an increase in the number of cells in the CSF. All patients exhibit accumulation of an unusual branched-chain fatty acid, phytanic acid (3,7,11,15-tetramethylhexadecanoic acid), in blood and tissues. Biochemically, the disease is caused by the deficiency of phytanoyl-CoA hydroxylase (PhyH), a peroxisomal protein catalyzing the first step in the alpha-oxidation of phytanic acid. We have purified PhyH from rat-liver peroxisomes and determined the N-terminal amino-acid sequence, as well as an additional internal amino-acid sequence obtained after Lys-C digestion of the purified protein. A search of the EST database with these partial amino-acid sequences led to the identification of the full-length human cDNA sequence encoding PhyH: the open reading frame encodes a 41.2-kD protein of 338 amino acids, which contains a cleavable peroxisomal targeting signal type 2 (PTS2). Sequence analysis of PHYH fibroblast cDNA from five patients with Refsum disease revealed distinct mutations, including a one-nucleotide deletion, a 111-nucleotide deletion and a point mutation. This analysis confirms our finding that Refsum disease is caused by a deficiency of Phy