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  • mitochondria  (53)
  • Springer  (53)
  • National Academy of Sciences
  • Periodicals Archive Online (PAO)
  • 1995-1999  (53)
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  • 1997  (53)
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  • Springer  (53)
  • National Academy of Sciences
  • Periodicals Archive Online (PAO)
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  • 1995-1999  (53)
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  • 1
    ISSN: 1573-4919
    Keywords: muscular diseases ; mitochondria ; MTDNA ; ATP synthase ; human
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The expression of several mitochondrial and nuclear genes involved in ATP production was examined in cells cultured from muscle biopsies of patients harboring mitochondrial pathologies. The transcript patterns in muscle cells from the patients affected by carnitine palmitoyl transferase II or 2-ketoglutarate dehydrogenase deficiencies were almost similar to control patterns. In the opposite, patterns were strikingly abnormal in all the other cell cultures from patients with defects in enzymatic complexes involved in oxidative phosphorylation: mitochondrial complex II and III deficiencies, two MELAS syndromes (myopathy, encephalopathy, lactic acidosis and stroke like episodes), a case of Kearns-Sayre syndrome and a case of chronic progressive external ophthalmoplegia. In cultured muscle cells from patients with mtDNA mutations, the percentage of mutated mtDNA was low as compared with those determined in the corresponding skeletal muscle biopsy. Moreover, the complex II defect resulting of a nuclear mutation was not expressed in the cell cultures. Thus, an undetermined transcriptional event, transmitted from muscle biopsies to cultured muscle cells, should be involved to account for such abnormal transcript patterns.
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  • 2
    ISSN: 1573-4919
    Keywords: porphyrin derivative ; mitochondria ; ascites ; singlet oxygen ; photosensitization ; lipid peroxidation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract With a view to locate porphyrins for use in photodynamic therapy (PDT), the new modality of cancer treatment we have evaluated the ability of a novel water soluble porphyrin meso-tetrakis[4-(carboxymethyleneoxy)phenyl]porphyrin (T4CPP) to induce damage to mitochondria during photosensitization. T4CPP, when exposed to visible light, induced lipid peroxidation in rat liver mitochondria as assessed by the formation of thiobarbituric acid reactive substances (TBARS), conjugated dienes (CD) and lipid hydroperoxides (LOOH). The effect on mitochondrial function was assessed by estimating the activity of succinate dehydrogenase (SDH). The peroxidation induced was observed to be time- and concentration- dependent. Analysis of product formation and selective inhibition by scavengers of reactive oxygen species showed that the oxidative damage observed was mainly due to singlet oxygen (1O2) and partly due to other reactive species. T4CPP plus light also caused significant lipid peroxidation in Sarcoma 180 ascites tumour mitochondria. Our studies indicate that T4CPP has the potential to photoinduce damage in hepatic and ascites mitochondria, a crucial site of damage in PDT. (Mol Cell Biochem 166: 25-33, 1997)
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  • 3
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    Molecular and cellular biochemistry 174 (1997), S. 325-328 
    ISSN: 1573-4919
    Keywords: ageing ; theory ; mitochondria ; respiratory chain ; mitochondrial DNA mutations
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Mitochondria are believed to be involved in human ageing. Whilst it is clear that various mitochondrial DNA mutations do accumulate in human tissues with age, whether or not they interfere with respiratory chain function is uncertain. We question the results of previous studies which have measured respiratory chain function in human skeletal muscle with age. Whilst cytochrome c oxidase deficient fibres are a real finding in skeletal muscle, the contribution of mitochondrial DNA mutations to human ageing is still controversial. Our results show for mitochondria to be involved in ageing then it must be through a more subtle mechanism than a global decline in respiratory chain function. (Mol Cell Biochem 174: 325–328, 1997)
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  • 4
    ISSN: 1573-4919
    Keywords: outlet obstruction ; bladder ; mitochondria ; transcription ; RNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Using the rabbit model, we showed that partial outlet obstruction of theurinary bladder causes significant changes in the status and expression ofthe mitochondrial (mt) genetic system in bladder smooth muscle immediatelyafter obstruction is initiated. Here we investigate quantitatively theseverity of the mt genetic response to partial outlet obstruction in bothshort- and long-term obstructed rabbits. Based on previous functionalstudies, bladders with mass 〈 6 fold greater than control were consideredcompensated; bladders with mass 〉 6 fold that of control were considereddecompensated. Analyses of DNA from compensated rabbit bladders showed thatrelative mt genome copy number decreased to 30% of control values.Transcript analyses for these samples showed that mt RNA levels increased 3fold to compensate for lower template copy number. Analysis of decompensatedbladders demonstrated that mt genome copy number increased to approximately90% of control levels; mt transcripts progressively decreased inthese samples by as much as 30 fold. In contrast, transcription of amt-related nuclear gene decreased 3-9 fold in compensated bladders butincreased 10-30 fold in decompensated bladders. Activity for the cytochromeoxidase complex, and for the mt enzyme citrate synthase, decreased steadilywith increasing bladder hypertrophy. These data suggest that bladderdysfunction following partial outlet obstruction is mediated partly by asignificant loss in mt and mt-related nuclear gene coordination.
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  • 5
    ISSN: 1573-4919
    Keywords: calorimetry ; cardiac muscle ; mitochondria ; oxidative phosphorylation ; atractyloside ; dinitrophenol ; ectonucleotidase ; respiratory control
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract A novel flow calorimetric technique was developed to study the energy turnover of myocardial mitochondria. Cylindrical strands of cardiac muscle (trabeculae) weighing 100–500 µg were isolated from guinea-pig heart and mounted in a tubular recording chamber which was continuously perfused with physiological salt solution at 37°C. The temperature difference between the upstream and the downstream side of the chamber, which is proportional to the rate of heat production of the trabecula, was measured at high resolution. In this way the rate of energy expenditure of isolated cardiac muscle could be recorded continuously for several hours. When the preparations were superfused with an 'intracellular' solution containing 5 mM pyruvate and 2 mM malate as substrates, permeabilization of the sarcolemma with 25 µM digitonin induced a marked increase in the measured heat rate in the presence of 2 mM ADP. The major fraction of the ADP sensitive heat production (83%) could be blocked with 400 µM at ractyloside, an inhibitor of the adeninenucleotide translocase, and by 600 µM α-cyano-4-hydroxycinnamate, an inhibitor of monocarboxylate/H+ co-transport. The atractyloside sensitive heat production was abolished in anoxic solution. These results suggest that the atractyloside-sensitive heat production (21.8 ± 3.5 mW cm-3 of tissue) was attributable to oxidative phosphorylation. The mitochondria apparently remained intact after treatment with digitonin, since application of the uncoupler 2,4-dinitrophenol (DNP) produced a very large increase in heat rate. A minor fraction of the heat rate induced by ADP in permeabilized cardiac muscle preparations (17%) was not sensitive to atractyloside. This component was also seen before application of digitonin and was probably related to ectonucleotidases. In conclusion, our calorimetric technique allows investigation of the energy metabolism of myocardial mitochondria 'in situ', i.e. without destroying the microarchitecture of cardiac muscle cells. (Mol Cell Biochem 174: 101–113, 1997)
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  • 6
    ISSN: 1573-4919
    Keywords: mitochondria ; cyclosporin ; cyclophilin ; channels ; permeability
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Mitochondria possess an inner membrane channel, the permeability transition pore, which is inhibited by cyclosporin A (CBA) and by matrix protons. As suggested recently by our laboratory, pore closure by these inhibitors may be due to dissociation of mitochondrial cyclophilin (CyP-M), a matrix peptidyl-prolyl-cis-trans isomerase, from its putative binding site on the pore. Unbinding of CyP-M would follow a CsA-dependent or proton-dependent change in conformation of the CyP-M molecule. It is interesting that upon binding of CsA the enzymatic activity of CyP-M is inhibited, but it is not clear whether this event plays a role in pore inhibition. Here we report experiments designed to further test the role of CyP-M in pore function. Our results indicate that CyP-M-dependent and independent mechanisms of pore activation may exist, and that the peptidylprolyl-cis-trans-isomerase activity of CyP-M is not necessarily involved in pore modulation by CyP-M. (Mol Cell Biochem 174: 181–184, 1997)
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  • 7
    ISSN: 1573-4919
    Keywords: mitochondria ; calcium ; permeability transition ; vasopressin ; glucagon ; thapsigargin ; protein kineses and phosphatases ; rat hepatocytes
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Ca2+ functions as an intracellular signal to transfer hormonal messages to different cellular compartments, including mitochondria, where it activates intramitochondrial Ca2+-dependent enzymes. However, excessive mitochondrial Ca2+ uptake can promote the mitochondrial permeability transition (MPT), a process known to be associated with cell injury. The factors controlling mitochondrial Ca2+ uptake and release in intact cells are poorly understood. In this paper, we investigate mitochondrial Ca2+ accumulation in intact hepatocytes in response to the elevation of cytosolic Ca2+ levels ([Ca2+]c) induced either by a hormonal stimulus (vasopressin), or by thapsigargin, an inhibitor of the endoplasmic reticulum Ca2+ pump. After stimulation, cells were rapidly permeabilized for the determination of the mitochondrial Ca2+ content (Ca2+_m) and to analyze the susceptibility of the mitochondria to undergo the MPT. Despite very similar levels of [Ca2+]c elevation, vasopressin and thapsigargin had markedly different effects on mitochondrial Ca2+ accumulation. Vasopressin caused a rapid (〈 90 sec), but modest (〈 2 fold) increase in Ca2+m that was not further increased during prolonged incubations, despite a sustained [Ca2+]c elevation. By contrast, thapsigargin induced a net Ca2+ accumulation in mitochondria that continued for up to 30 min and reached Ca2+_m levels 10–20 fold over basal. Accumulation of mitochondrial Ca2+ was accompanied by a markedly increased susceptibility to undergo the MPT. Both mitochondrial Ca2+ accumulation and MPT activation were modulated by treatment of the cells with inhibitors of protein kineses and phosphatases. The results indicate that net mitochondrial Ca2+ uptake in response to hormonal stimulation is regulated by processes that depend on protein kinase activation. These controls are inoperative when the cytosol is flooded by Ca2+ through artificial means, enabling mitochondria to function as a Ca2+ sink under these conditions. (Mol Cell Biochem 174: 173–179, 1997)
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  • 8
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    Molecular and cellular biochemistry 174 (1997), S. 215-219 
    ISSN: 1573-4919
    Keywords: MERRF ; mitochondria ; mtDNA ; genetics ; tRNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract MERRF (myoclonic epilepsy with ragged-red fibers) is a severe, multisystem disorder characterized by myoclonus, seizures, progressive cerebellar syndrome, muscle weakness, and the presence of ragged-red fibers in the muscle biopsy. MERRF is associated with heteroplasmic point mutations, either A8344G or T8356C, in the gene encoding the mitochondrial tRNALys. The human ro cell system was utilized to examine the phenotypic consequences of these mutations, and to investigate their molecular genetic causes. Wild-type and mutant transmitochondrial cell lines harboring a pathogenic point mutation at either A8344G or T8356C in the human mitochondrial tRNALys gene were isolated and examined. Mitochondrial transformants containing 100% mutated mitochondrial DNAs (mtDNAs) exhibited severe defects in respiratory chain activity, in the rates of protein synthesis, and in the steady-state levels of mitochondrial translation products as compared with mitochondrial transformants containing 100% wild-type mtDNAs. In addition, both mutant cell lines exhibited the presence of aberrant mitochondrial translation products. These results demonstrate that two different mtDNA point mutations in tRNALys result in fundamentally identical defects at the cellular level, and that these specific protein synthesis abnormalities contribute to the pathogenesis of MERRF. (Mol Cell Biochem 174: 215–219, 1997)
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  • 9
    ISSN: 1573-4919
    Keywords: diabetes ; carbon tetrachloride ; liver toxicity ; glutathione ; mitochondria ; Schisandra chinensis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The streptozotocin-induced short-term (2 week) diabetic rats showed an increase in susceptibility to carbon tetrachloride (CCl4)-induced hepatocellular damage. This diabetes-induced change was associated with a marked impairment in the hepatic glutathione antioxidant/detoxification response to CCl4 challenge, as indicated by the abrogation of the increases in hepatic reduced glutathione (GSH) level, glucose-6-phosphate dehydrogenase and microsomal glutathione S-transferases (GST) activities upon challenge with increasing doses of CCl4. While the hepatic GSH level was increased in diabetic rats, the hepatic mitochondrial GSH level and Se-glutathione peroxidase activity were significantly reduced. Insulin treatment could reverse most of the biochemical alterations induced by diabetes. Both insulin and schisandrin B (Sch B) pretreatments protected against the CCl4 hepatotoxicity in diabetic rats. The hepatoprotection was associated with improvement in hepatic glutathione redox status in both cytosolic and mitochondrial compartments, as well as the increases in hepatic ascorbic acid level and microsomal GST activity. The ensemble of results suggests that the diabetes-induced impairment in hepatic mitochondrial glutathione redox status may at least in part be attributed to the enhanced susceptibility to CCl4 hepatotoxicity. Sch B may be a useful hepatoprotective agent against xenobiotics-induced toxicity under the diabetic conditions. (Mol Cell Biochem 175: 225–232, 1997)
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  • 10
    ISSN: 1573-4919
    Keywords: heart ; vascular endothelium ; vascular smooth muscle ; confocal microscopy ; pH ; calcium ; sodium ; voltage probe ; heart ; endothelin-1 ; Angiotensin II ; PAF ; nucleus ; mitochondria ; SR ; cardiomyopathy ; cells interaction ; R-type Ca2+ channel ; excitation-contraction coupling ; dystrophic mouse
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract In recent years, fluorescence microscopy imaging has become an important tool for studying cell structure and function. This non invasive technique permits characterization, localisation and qualitative quantification of free ions, messengers, pH, voltage and a pleiad of other molecules constituting living cells. In this paper, we present results using various commercially available fluorescent probes as well as some developed in our laboratory and discuss the advantages and limitations of these probes in confocal microscopy studies of the cardiovascular system.
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  • 11
    ISSN: 1573-4919
    Keywords: thiamine deficiency ; mitochondria ; energy metabolism ; necrosis ; neuroblastoma cells
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Culture of neuroblastoma cells in the presence of low thiamine concentration (6 nM) and of the transport inhibitor amprolium leads to the appearance of signs of necrosis: the chromatin condenses, the oxygen consumption decreases and is uncoupled, the mitochondrial cristae are disorganized, the thiamine diphosphate-dependent dehydrogenase activities are impaired. When 10 µM thiamine are added to these cells, the basal respiration increases, the coupled respiration is restored and mitochondrial morphology is recovered within 1 h. Addition of succinate, which is oxidized via a thiamine diphosphate-independent dehydrogenase, to digitonin-permeabilized cells immediately restores a coupled respiration. Our results suggest that the slowing of the citric acid cycle is the cause of the biochemical lesion induced by severe thiamine deficiency and that part of the mitochondria remain functional. (Mol Cell Biochem 174: 121–124, 1997)
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  • 12
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    Molecular and cellular biochemistry 174 (1997), S. 189-192 
    ISSN: 1573-4919
    Keywords: nitric oxide ; mitochondria ; inflammation ; respiration ; astrocytes ; cytochrome oxidase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Nitric oxide (NO) at high levels is cytotoxic, and may be involved in a range of inflammatory, neurodegenerative, and cardiovascular/ischaemic pathologies. The mechanism of NO-induced cytotoxicity is unclear. Recently we and others have found that low (nanomolar) levels of NO reversibly inhibit mitochondrial respiration by binding to the oxygen binding site of cytochrome oxidase in competition with oxygen. This raises the apparent Km for oxygen of mitochondrial respiration into the physiological range, potentially making respiration sensitive to the oxygen level. The NO inhibition of oxygen consumption was seen in isolated cytochrome oxidase, mitochondria, brain nerve terminals, and cultured cells. Cultured astrocytes activated to express the inducible form of NO synthase produced up to 1 µM NO and strongly inhibited their own cellular respiration rate. This respiratory inhibition was rapidly reversed by removing the NO, and was due to the inhibition of cytochrome oxidase. These results suggest that any cell producing high levels of NO will inhibit its own respiration and that of surrounding cells, and make the respiration rate sensitive to the oxygen level. This inhibition of energy metabolism may contribute to cytotoxity or cytostasis in some pathologies. (Mol Cell Biochem 174: 189–192, 1997)
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  • 13
    ISSN: 1573-4919
    Keywords: mitochondria ; mitochondrial myopathies ; oxidative phosphorylation ; principal component analysis (PCA) ; biplot
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The mitochondrial pathologies are a heterogeneous group of metabolic disorders that are characterized by anomalies of oxidative phosphorylation, especially in the respiratory chain. The diagnosis of these pathologies involves many investigations among which biochemical study is at present the main tool. However, the analysis of the results obtained during such study remains complex and often does not make it possible to conclude clearly if a patient is affected or not by a biochemical and/or bioenergetic deficiency. This arises from two main problems: 1. The determination of control values from the whole set of variable values (affected and unaffected people). 2. The small size of the population studied and the large number of variables collected which present a rather large variability. To cope with these problems, the principal component analysis method is applied to the results obtained during our biochemical studies. This analysis makes it possible for each respiratory chain complex, to distinguish clearly two subsets of the whole population (affected and unaffected people) as well as to detect the variables which are the most discriminative. (Mol Cell Biochem 174: 149–156, 1997)
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  • 14
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    Molecular and cellular biochemistry 175 (1997), S. 37-42 
    ISSN: 1573-4919
    Keywords: mitochondria ; peroxisomes ; fatty acids metabolism ; coenzyme A deficiency ; pantothenic acid
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Hepatic coenzyme A (CoA) plays an important role in cellular lipid metabolism. Because mitochondria and peroxisomes represent the two major subcellular sites of lipid metabolism, the present study was designed to investigate the specific impact of hepatic CoA deficiency on peroxisomal as well as mitochondrial β-oxidation of fatty acids. CoA deficiency (47% decrease in free CoA and 23% decrease in total CoA) was produced by maintaining weanling male Sprague-Dawley rats on a semipurified diet deficient in pantothenic acid (the precursor of CoA) for 5 weeks. Hepatic mitochondrial fatty acid oxidation of short-chain and long-chain fatty acids were not significantly different between control and CoA-deficient rats. Conversely, peroxisomal poxidation was significantly diminished (38% inhibition) in livers of CoA-deficient rats compared to control animals. Peroxisomal β-oxidation was restored to normal levels when hepatic CoA was replenished. It is postulated that since the role of hepatic mi tochondrial β-oxidation is energy production while peroxisomal β-oxidation acts mainly as a detoxification system, the mitochondrial pathway of β-oxidation is spared at the expense of the peroxisomal pathway when liver CoA plummets. The present study may offer an animal model to investigate mechanisms involved in peroxisomal diseases. (Mol Cell Biochem 175: 37–42, 1997)
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  • 15
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    Molecular and cellular biochemistry 174 (1997), S. 91-96 
    ISSN: 1573-4919
    Keywords: insulin ; mitochondria ; Krebs cycle ; pyruvate ; succinate
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Our previous studies of insulin action have led us to the finding that insulin acts specifically on the mitochondrial Krebs cycle to stimulate, by 30%, the oxidation of carbons 2 and 3 of pyruvate to CO2. Insulin also stimulates the oxidation of both carbons of acetate. These carbons can be converted to CO2 only after passing through all of the reactions of the Krebs cycle more than once. Carboxyl groups, such as number 1 of pyruvate, are oxidized to CO2 without any effect of insulin, and can be converted to CO2 by extramitochondrial enzyme. We conclude that insulin must act on the complete intramitochondrial cycle and not on the four enzymes of the Krebs cycle which are present in the cytoplasm. The path taken by those carbons affected by insulin is traced through the complete Krebs cycle, and the necessity for this effect to be mitochondrial has been verified by demonstration of the same specific effect of insulin on the oxidation of the 2 and 3 carbons of succinate. The use of this phenomenon is proposed for the study not only of human diabetes, but of all mitochondrial disorders, by using 14C specifically labeled tracers in culture or biopsy material, or 13C labeled tracer material in vivo. (Mol Cell Biochem 174: 91–96, 1997)
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  • 16
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    Molecular and cellular biochemistry 174 (1997), S. 255-259 
    ISSN: 1573-4919
    Keywords: mitochondria ; neuroleptics ; oxidative phosphorylation ; complex I
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract There is increasing evidence that a defect of the mitochondrial respiratory chain is implicated in the development of Parkinson disease. Decreased complex I activity of the mitochondrial respiratory chain has been reported in platelets, muscle, and brain of patients with Parkinson disease. Extrapyramidal symptoms (e.g. parkinsonism and dystonic reactions) are major limiting side effects of neuroleptics. Experimental evidence suggests that neuroleptics inhibit complex I in rat brain. There has not been a study of the effects of neuroleptics in human tissue, however. We therefore analyzed the activities of complexes I + III, complexes II + III, succinate dehydrogenase, complex IV (cytochrome c oxidase), and of citrate synthase in normal human brain cortex after the addition of haloperidol and chlorpromazine and the atypical neuroleptics risperidone, zotepine, and clozapine. Activity of complex I was progressively inhibited by all neuroleptics. Half maximal inhibition (IC50) was 0.1 mM fo r haloperidol, 0.4 mM for chlorpromazine, and 0.5 mM for risperidone and zotepine. Clozapine had no effect on enzyme activity at concentrations up to 0.5 mM, followed by a slow decline with a maximum inhibition of 70% at 10 mM. IC50 was at about 2.5 mM. Thus, the concentration of clozapine needed to cause 50% inhibition of the activity of complexes I and III was about 5 times that of zotepine and risperidone, about 6 times that of chlorpromazine, and 25 times that of haloperidol. The inhibition thus paralleled the incidence of extrapyramidal effects caused by the different neuroleptics as they are known from numerous clinical studies. Our data support the hypothesis that neuroleptic-induced extrapyramidal side effects may be due to inhibition of the mitochondrial respiratory chain. (Mol Cell Biochem 174: 255–259, 1997)
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  • 17
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    Molecular and cellular biochemistry 174 (1997), S. 277-281 
    ISSN: 1573-4919
    Keywords: mitochondria ; myopathy ; inclusion body myositis ; neuropathy ; vasculitis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract Computer retrieval in a database, comprising 7,225 muscle cases, revealed that mitochondrial myopathies do not occur more frequently in inflammatory myopathies (3.74%) than in the whole series (3.69%). A more detailed study of inclusion body myositis (IBM), however, showed that severe mitochondrial alterations were apparent in about twice as many IBM cases as expected. This confirms recent studies of others although a causal relationship has thus far not been established. Identification of mitochondrial deletions by Southern blotting corresponded to the presence of severe structural abnormalities of mitochondria. Peripheral neuropathy of variable severity was noted in all cases of IBM and mitochondrial myopathy. By contrast, the association of severe mitochondrial abnormalities with polymyositis, systemic scleroderma, and vasculitis observed in some cases of the present series may be incidental or age dependent. (Mol Cell Biochem 174: 277–281, 1997)
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  • 18
    ISSN: 1573-4919
    Keywords: mercury ; rat kidney ; mitochondria ; oxidative phosphorylation ; FoF1-ATPase ; ATP synthesis ; ATP hydrolysis ; oxidative stress
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Medicine
    Notes: Abstract The effects of Hg(II) on bioenergetic and oxidative status of rat renal cortex mitochondria were evaluated both in vitro, and in vivo 1 and 24 h after treatment of animals with 5 mg HgCl2/kg ip. The parameters assessed were mitochondrial respiration, ATP synthesis and hydrolysis, glutathione content, lipid peroxidation, protein oxidation, and activity of antioxidant enzymes. At low concentration (5 µM) and during a short incubation time, Hg(II) uncoupled oxidative phosphorylation while at slightly higher concentration or longer incubation time the ion impaired the respiratory chain. The rate of ATP synthesis and the phosphorylation potential of mitochondria were depressed, although inhibition of ATP synthesis did not exceed 50%. In vivo, respiration and ATP synthesis were not affected 1 h post-treatment, but were markedly depressed 24 h later. ATP hydrolysis by submitochondrial particle FoF1-ATPase was inhibited (also by no more than 50%) both in vitro, and in vivo 1 and 24 h post-treatment. Hg(II) induced maximum ATPase inhibition at about 1 uM concentration but did not have a strong inhibitory effect in the presence of Triton X-100. Oxidative stress was not observed in mitochondria 1 h post-treatment. However, 24 h later Hg(II) reduced the GSH/GSSG ratio and increased mitochondrial lipid peroxidation and protein oxidation, as well as inhibited GSH-peroxidase and GSSG-reductase activities. These results suggest that the following sequence of events may be involved in Hg(II) toxicity in the kidney: (1) inhibition of FoFl-ATPase, (2) uncoupling of oxidative phosphorylation, (3) oxidative stress-associated impairment of the respiratory chain, and (4) inhibition of ATP synthesis.
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  • 19
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    Bioscience reports 17 (1997), S. 17-21 
    ISSN: 1573-4935
    Keywords: Hyperthyroidism ; mitochondria ; uncoupling ; proton-leak ; pump-slip
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract During the past years many efforts have been made to elucidate the origin of the uncoupling mechanisms induced by hyperthyroidism in mitochondria. Two main mechanisms have been proposed: a classical protonophoric uncoupler mechanism, considering the action of thyroid hormones at the level of the lipid membrane bilayer, and a slipping mechanism with more localized effects at the level of the redox proton pumps. This short review is devoted to comparing and discussing the evidence against and in favour of these two mechanisms.
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  • 20
    ISSN: 1573-4935
    Keywords: Calcium ; cyclosporin A ; lipid peroxidation ; mitochondria ; mitochondrial membrane permeability transition ; protein oxidation ; reactive oxygen species
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract We have provided evidence that mitochondrial membrane permeability transition induced by inorganic phosphate, uncouplers or prooxidants such as t-butyl hydroperoxide and diamide is caused by a Ca2+-stimulated production of reactive oxygen species (ROS) by the respiratory chain, at the level of the coenzyme Q. The ROS attack to membrane protein thiols produces cross-linkage reactions, that may open membrane pores upon Ca2+ binding. Studies with submitochondrial particles have demonstrated that the binding of Ca2+ to these particles (possibly to cardiolipin) induces lipid lateral phase separation detected by electron paramagnetic resonance experiments exploying stearic acids spin labels. This condition leads to a disorganization of respiratory chain components, favoring ROS production and consequent protein and lipid oxidation.
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  • 21
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    Bioscience reports 17 (1997), S. 67-76 
    ISSN: 1573-4935
    Keywords: Apoptosis ; necrosis ; mitochondria ; megachannel ; permeability transition ; programmed cell death ; poteases
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Mitochondrial permeability transition (PT) involves the formation of proteaceous, regulated pores, probably by apposition of inner and outer mitochondrial membrane proteins which cooperate to form the mitochondrial megachannel (=mitochondrial PT pore). PT has important metabolic consequences, namely the collapse of the mitochondrial transmembrane potential, uncoupling of the respiratory chain, hyperproduction of superoxide anions, disruption of mitochondrial biogenesis, outflow of matrix calcium and glutathione, and release of soluble intermembrane proteins. Recent evidence suggests that PT is a critical, rate limiting event of apoptosis (programmed cell death): (i) induction of PT suffices to cause apoptosis; (ii) one of the immediate consequences of PT, disruption of the mitochondrial transmembrane potential (ΔΨm), is a constant feature of early apoptosis; (iii) prevention of PT impedes the ΔΨm collapse as well as all other features of apoptosis at the levels of the cytoplasma, the nucleus, and the plasma membrane; (iv) PT is modulated by members of the apoptosis-regulatory bcl-2 gene family. Recent data suggest that the acquisition of the apoptotic phenotype, including characteristic changes in nuclear morphology and biochemistry (chromatin condensation and DNA fragmentation), depends on the action of apoptogenic proteins released from the mitochondrial intermembrane space.
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  • 22
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    Bioscience reports 17 (1997), S. 251-257 
    ISSN: 1573-4935
    Keywords: Leak ; mitochondria ; proton
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract The proton conductance of the mitochondrial inner membrane can be quantified by applying Ohm's law to the experimentally determined protonmotive force and the proton current flowing around the proton circuit in the absence of ATP synthesis or ion transport. This last parameter is derived from the rate of State 4 respiration multiplied by the H+/O stoichiometry for the substrate. When the activity of the dehydrogenase supplying electrons to the respiratory chain is progressively increased the proton conductance increases rapidly when the protonmotive force is greater than 220 mV. The consequences of this non-ohmic relationship are discussed.
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    Bioscience reports 17 (1997), S. 259-272 
    ISSN: 1573-4935
    Keywords: Superoxide ; hydrogen peroxide ; electron leak ; proton leak ; H+/2e ; membrane potential ; reactive oxygen cycle ; heat production ; mitochondria
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Recently, we proposed a hypothetical model of coexistence of “Reactive oxygen cycle” with Q cycle and H+ cycle in mitochondrial respiratory chain to combine both processes of univalent electron leak for production of superoxide and of proton leak across inner mitochondrial membrane. This review presents a more detailed description of this model and summarizes the supporting experimental evidence obtained.
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  • 24
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    Bioscience reports 17 (1997), S. 347-366 
    ISSN: 1573-4935
    Keywords: Reactive oxygen species ; mitochondria ; pore ; apoptosis ; uncoupling ; non-coupled respiration ; aconitase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract New facts and ideas concerning the membrane-linked mechanisms preventing superoxide formation are summarised here. It is assumed that aerobic cells possess several lines of anti-ROS defence, including optimisation of the intracellular oxygen concentration, decrease in the concentration and life-time of one-electron O2 reductants such as CoQH; and mitochondrial and cell selections, i.e. elimination of mitochondria and cells producing ROS at high rate. It is postulated that ROS-dependent pore opening and ROS-dependent apoptosis are involved in mitochondrial and cell selections.
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  • 25
    ISSN: 1573-5028
    Keywords: antiproliferation ; Arabidopsis ; mitochondria ; subcellular fractionation ; tobacco
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    Topics: Biology
    Notes: Abstract This report describes the cloning and characterization of a plant cDNA coding for a protein which shows high amino acid sequence similarity with prohibitin, whose gene is associated with antiproliferative activity in mammalian cells. Arabidopsis thaliana and Nicotiana tabacum prohibitin complete cDNAs were isolated, and the expression pattern of prohibitin was examined using polyclonal antibodies raised against the Arabidopsis recombinant prohibitin expressed in Escherichia coli. A single immunoreactive protein was detected in various plant species and in all Arabidopsis organs examined. Subcellular fractionation using tobacco leaves revealed prohibitin in a mitochondrial-enriched fraction. Phylogenetic conservation of prohibitin's amino acid sequence and subcellular localization suggests a similar function in plants, yeast and mammals.
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  • 26
    ISSN: 1573-5028
    Keywords: Betula pendula Roth ; differential display ; gene expression ; mitochondria ; oxidative stress ; ozone ; phosphate translocator
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    Topics: Biology
    Notes: Abstract We have isolated by DDRT-PCR (differential-display reverse-transcription polymerase chain reaction) and cDNA library screening a 1.3 kb cDNA corresponding to a strongly ozone-inducible transcript from birch (Betula pendula Roth). Nucleotide sequence analysis suggests that it encodes a mitochondrial phosphate translocator protein (Pi c), the first one isolated from plants. The isolated birch mitochondrial phosphate translocator cDNA (designated Mpt1) contains an open reading frame of 1092 bases encoding a 364 amino acid polypeptide. The deduced protein is 66% similar to bovine Pic isoform B. Comparison of the N-terminal amino acid sequence to known mammalian Pic proteins and the existence of an in-frame stop codon upstream of the initiation codon suggest that the isolated cDNA is full-length. Southern hybridization analysis of birch genomic DNA shows that Mpt1 is a single-copy gene. Accumulation of Mpt1 mRNA during oxidative stress imposed by ozone is detectable already at 2 h and it is at maximum ca. 12 h after the beginning of an 8 h ozone exposure (150 ppb). A second O3 peak at 48–56 h did not increase transcript levels further. O3 exposure for 2 h was sufficient for Mpt1 induction. Birch Mpt1 transcript levels remain at moderately low level during leaf development and is lower in roots and leaves when compared to young shoots undergoing wood formation and lignification.
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  • 27
    ISSN: 1573-5028
    Keywords: Arabidopsis thaliana/ ; carotenoid ; crtE/ ; gene cloning ; GGPP synthase ; mitochondria
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    Topics: Biology
    Notes: Abstract We have cloned a new geranylgeranyl pyrophosphate (GGPP) synthase gene, designated GGPS6/, from Arabidopsis thaliana genomic DNA. Nucleotide sequence analysis revealed that the GGPS6 gene contains an open reading frame coding for a protein of 343 amino acid residues with a calculated molecular mass of 37 507 Da. Also, the gene is not interrupted by an intron. The predicted amino acid sequence of the GGPS6 gene shows significant homology (34.0–57.7%) with other GGPP synthases from Arabidopsis. The GGPS6 protein contains a N-terminal signal peptide which is thought to function as an organelle targeting sequence. In fact, the GGPS6-GFP fusion protein was found to be localized exclusively in mitochondria when expressed in tobacco BY-2 cells. In vitro analysis of the enzyme activity as well as genetic complementation analysis with Erwinia uredovora crt gene cluster expressed in Escherichia coli showed that the GGPS6 gene most certainly encodes a GGPP synthase catalyzing the conversion of farnesyl pyrophosphate to GGPP.
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  • 28
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    Bioscience reports 17 (1997), S. 273-279 
    ISSN: 1573-4935
    Keywords: Uncoupling ; mitochondria ; free radicals ; thyroid hormones ; steroid hormones
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    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Recently, it was proposed that the thyroid hormone-mediated uncoupling in mitochondria is involved in the cellular defence system against free radicals (Skulachev V.P. (1996) Quart. Rev. Biophys. 29:169–202). This phenomenon was named “mild” uncoupling. It was postulated to be a protein-mediated process controlled by several factors. The data reported during the past 40 years, pointing to the protein-mediated uncoupling mechanism in mitochondria, are reviewed in a context of hypothetical properties of “mild” uncoupling. The mechanism of “mild” uncoupling is suggested to be the following: (a) mitochondria possess protein(s) that regulate the proton permeability of inner mitochondrial membrane; (b) these proteins are regulated by binding of an unidentified low-molecular-weight endogenous compound with properties resembling those of the most active artificial uncouplers like FCCP and SF6847; (c) the interaction of this compound with its target protein(s) is modulated by a thyroid hormone in a positive (i.e. enhancing the proton permeability) way and by sex steroid hormones in a negative way; (e) endogenous fatty acids can attenuate the influence of both thyroid and steroid hormones.
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  • 29
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    Bioscience reports 17 (1997), S. 319-333 
    ISSN: 1573-4935
    Keywords: Plant alternative oxidase ; mitochondria ; oxidative stress ; active oxygen species
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    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Current understanding of the structure and function of the plant alternative oxidase is reviewed. In particular, the role of the oxidase in the protection of tissues against oxidative stress is developed.
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    Bioscience reports 17 (1997), S. 521-527 
    ISSN: 1573-4935
    Keywords: Glyceraldehyde-3-phosphate dehydrogenase ; glycolysis ; oxidation ; uncoupling ; thiols ; hydrogen peroxide ; mitochondria
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    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract Data on alterations of the properties of glyceraldehyde-3-phosphate dehydrogenase upon oxidation of its functional groups are reviewed; a mechanism of uncoupling of oxidation and phosphorylation in glycolysis is considered. Possible ways of regulating uncoupling, and the physiological importance of this process, are discussed.
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  • 31
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    Bioscience reports 17 (1997), S. 557-567 
    ISSN: 1573-4935
    Keywords: Reactive oxygen species (ROS) ; mitochondria ; Ca2+ transport ; energy transduction ; electron paramagnetic resonance (EPR)
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    Topics: Biology , Chemistry and Pharmacology
    Notes: Abstract The relationship between Ca2+ transport and energy transduction of myocardial mitochondria in the presence of reactive oxygen species was investigated. Following treatment with oxygen free radicals [superoxide(O 2 • ) or hydroxyl radical (•)OH], lipid free radicals in myocardial mitochondrial membrane could be detected by using the method of EPR spin trap. Simultaneously there were obvious alterations in the free Ca2+ ([Ca2+]m) in the mitochondrial matrix; the physical state of membrane lipid; the efficiency of oxidative phosphorylation (ADP/O); the value of the respiratory control ratio (RCR); and the membrane potential of the inner membrane of myocardial mitochondria. If the concentrations of reactive oxygen species were reduced by about 30%, the alterations in the physical state of the membrane lipid and energy transduction of myocardial mitochondria were not observed, but the changes in Ca2+ homeostasis remained. We conclude that Ca2+ transport by myocardial mitochondria is more sensitive to agents such as (O 2 • ) or •OH, etc. than are oxidation phosphorylation and the respiratory chain.
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  • 32
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    Plant and soil 191 (1997), S. 189-203 
    ISSN: 1573-5036
    Keywords: bacteroids ; leghaemoglobin ; mitochondria ; nitrogenase ; oxygen ; pro-plastids ; regulation ; respiration
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Respiration and nitrogen fixation in legume root nodules is considered to be limited by the rate at which O2 from the atmosphere can enter nodules. A thin diffusion barrier in the inner cortex, restricts access to the central tissue where there is a high demand for and low concentration of O2. Observed variations in rates of nodule activities in response to imposed stresses, are often attributed to variations in the diffusion resistance of the barrier. In the present work, alternative or supplementary metabolic mechanisms are considered. Aspects of nodule structure and of metabolism underlying nodule activities are reviewed in terms of components of the symbiotic system, the nature of steady states and in relation to homeostasis of low concentration of O2 within the bacteroid-filled host cells. It is suggested that variations in O2-demand of both mitochondria and bacteroids, serve to preserve nitrogenase activity by poising O2 concentration within ‘safe’ limits. Further, data from isolated soybean bacteroids suggest that nitrogenase is converted to a less active but more robust form, in the presence of O2 in excess of about 70 nM, thus protecting nitrogenase from irreversible inactivation by excess O2. This regulation is rapidly-reversible when O2 concentration falls below about 0.1 µM. Respiration by large numbers of host mitochondria in the periphery of infected nodule cells, adjacent to gas-filled intercellular spaces, is considered to play an important part in maintaining a steep gradient of O2 concentration in this zone. Also, it is possible that variations in nodule O2 demand may be involved in the apparent variations in resistance of the diffusion barrier. It is concluded that there are many biochemical components which should be considered, along with possible changes to the diffusion barrier, when the effects of imposed stresses on nodule activities are being analysed.
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    Fish physiology and biochemistry 16 (1997), S. 531-541 
    ISSN: 1573-5168
    Keywords: mitochondria ; muscle ; thermal acclimatization ; fish ; substrate preferences ; phospholipid fatty acid composition ; oxidative capacity
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    Topics: Biology
    Notes: Abstract Cold-acclimation of rainbow trout brings only limited changes in muscle metabolic capacities, but marked modifications in membrane composition. Thus, we examined whether the functional properties of mitochondria from trout red muscle were modified by seasonal temperature acclimatization. Mitochondria from fall-acclimatized trout had higher maximal capacities (state 3 rates) for the oxidation of pyruvate and acyl carnitines at 12 and 20 °C than mitochondria isolated from summer-acclimatized trout. For these substrates, the increased oxidative capacity completely compensated for the seasonal drop in temperature. Pyruvate and palmitoyl carnitine were consistently the preferred substrates, while decanoyl and octanoyl carnitine were oxidized at higher rates than glutamine, particularly in fall trout. State 4 rates of oxygen uptake (obtained when all ADP has been converted to ATP) differed less among substrates, but varied seasonally. State 4 rates at 12 and 20 °C were higher in mitochondria isolated from fall than summer trout. At low temperatures, the Q10 of both maximal and state 4 rates of substrate oxidation tended to be higher for mitochondria from fall trout. The apparent Arrhenius activation energy (Ea) for mitochondrial pyruvate oxidation was higher in fall than summer trout whereas the Ea’s for palmitoyl carnitine and decanoyl carnitine oxidation did not change. The fatty acids of mitochondrial phospholipids from fall trout were more polyunsaturated than those from summer trout, with 12% more double bonds occurring than in summer trout, suggesting that membrane restructuring may be involved in the observed compensatory responses.
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  • 34
    ISSN: 1573-6881
    Keywords: bc 1 complex ; mitochondria ; cytochromes ; transmembrane pH difference ; H+/e − ratio ; decoupling ; azide ; arachidonate
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    Topics: Biology , Chemistry and Pharmacology , Physics
    Notes: Abstract The effect of different anions on the steady-state proton translocation in bovine bc 1 complex reconstituted in liposomes was studied. The H+/e − ratio for vectorial proton translocation is at the steady state definitely lower than that measured at level flow, (0.3 vs. 1.0). The presence of azide or arachidonate at micro- and submicromolar concentrations, respectively, gave a substantial reactivation of the proton pumping activity at the steady state, without any appreciable effect on respiration-dependent transmembrane pH difference. Addition of azide to turning-over bc 1 vesicles also caused a transition of b cytochromes toward oxidation. The results are discussed in terms of possible involvement of an acidic residue in the protonation of the semiquinone/quinol couple at the N side of the membrane.
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  • 35
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    Journal of bioenergetics and biomembranes 29 (1997), S. 109-119 
    ISSN: 1573-6881
    Keywords: ETS domain ; gene expression ; mammalian cells ; mitochondria ; nuclear respiratory factors ; oxidative phosphorylation ; regulation ; respiratory chain ; transcription
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    Topics: Biology , Chemistry and Pharmacology , Physics
    Notes: Abstract The majority of gene products required for mitochondrial respiratory function are encoded in the nuclear genome. These include most of the respiratory subunits and all of the proteins that regulate the mitochondrial genetic system. One approach to understanding nucleo-mitochondrial interactions in mammalian cells is to identify the nuclear transcription factors that are common to the expression of these gene products. This has led to the purification and molecular cloning of nuclear respiratory factors, NRF-1 and NRF-2. The DNA binding and transcriptional specificities of these proteins have implicated them in the expression of many respiratory subunits along with key components of the mitochondrial transcription, replication, and heme biosynthetic machinery. In addition, tissue-specific transcription factors have been linked to the coordinate synthesis of contractile proteins and muscle-specific respiratory subunits whereas other more ubiquitous factors may have a dual function in nuclear and mitochondrial gene activation. These findings provide a framework for further investigations of the nuclear genetic mechanisms that integrate the expression of the respiratory apparatus with that of other cellular systems during growth and development.
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  • 36
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    Keywords: Cardiolipin metabolism ; CCL16-B2 cells ; mitochondria
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    Topics: Biology , Chemistry and Pharmacology , Physics
    Notes: Abstract The metabolism of cardiolipin was investigated in a Chinese hamster lung fibroblast cell line CCL16-B2 deficient in oxidative energy metabolism and its parental cell line CCL16-B1. Mitochondrial enzyme activities involved in de novo cardiolipin biosynthesis were elevated in CCL16-B2 cells compared with CCL16-B1 cells, indicating initially an elevation in cardiolipin biosynthesis. Content of all phospholipids, including cardiolipin and its precursors, and high energy nucleotides were unaltered in CCL 16-B2 cells compared to CCL 16-B1 cells. When cells were incubated with [1,3-3H]glycerol for up to 4 h radioactivity incorporated into cardiolipin in CCL16-B2 cells did not differ compared with CCL16-B1 cells. In contrast, radioactivity incorporated into phosphatidylglycerol, the immediate precursor of cardiolipin, was elevated over 2-fold in CCL16-B2 cells compared with CCL16-B1 cells. Analysis of the fatty acid molecular species in cardiolipin revealed alterations in the level of unsaturated but not saturated fatty acids in B2 compared with B1 cells. In vivo cardiolipin remodeling, that is, the deacylation of cardiolipin to monolysocardiolipin followed by reacylation back to cardiolipin, with [1-14C]palmitate and [l-14C]oleate and in vitro mitochondrial phospholipid remodeling with [1-14C]linoleate were altered in CCL16-B2 cells compared to CCL16-B1 cells. Since both the appropriate content and molecular composition of cardiolipin is required for optimum mitochondrial oxidative phosphorylation, we suggest that the difference in CL molecular species composition observed in CCL16-B2 cells, mediated by alterations in in vivo cardiolipin remodeling, may be one of the underlying mechanisms for the reduction in oxidative energy production in CCL16-B2 cells.
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    Journal of bioenergetics and biomembranes 29 (1997), S. 525-531 
    ISSN: 1573-6881
    Keywords: Porin ; ion channel ; mitochondria ; VDAC ; electron microscopy ; sequence analysis ; β-barrel
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    Topics: Biology , Chemistry and Pharmacology , Physics
    Notes: Abstract There is considerable evidence that the voltage-gated mitochondrial channel VDAC forms a β-barrel pore. Inferences about the number and tilt of β-strands can be drawn from comparisons with bacterial β-barrel pores whose structures have been determined by x-ray crystallography. A structural model for VDAC is proposed (based on sequence analysis and electron crystallography) in which the open state is like that of bacterial porins with several important differences. Because VDAC does not occur as close-packed trimers, there are probably fewer interpore contacts than in the bacterial porins. VDAC also appears to lack a large, fixed intraluminal segment and may not have as extensive a region of uniformly 35°-tilted β-strands as do the bacterial porins. These structural differences would be expected to render VDAC's β-barrel less stable than its bacterial counterparts, making major conformational changes like those associated with gating more energetically feasible. A possible gating mechanism is suggested in which movement of the N-terminal α-helix out of the lumen wall triggers larger-scale structural changes.
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  • 38
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    Journal of bioenergetics and biomembranes 29 (1997), S. 11-17 
    ISSN: 1573-6881
    Keywords: Protein targeting ; protein import ; mitochondria ; molecular chaperones
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    Topics: Biology , Chemistry and Pharmacology , Physics
    Notes: Abstract Protein import into mitochondria is initiated by the recognition and binding of precursor proteins by import components in the cytosol, on the mitochondrial surface, and in the mitochondrial outer membrane. Following their synthesis on cytoplasmic ribosomes, some precursor proteins interact with molecular chaperones in the cytosol which function in maintaining the precursor protein in an import-competent state and may also aid in the delivery of the precursor to the mitochondria. A multisubunit protein import receptor then recognises and binds precursor proteins before feeding them into the outer membrane import site. Some proteins are sorted from the import site into the outer membrane, but most precursor proteins travel through the outer membrane import site into the mitochondria, where the later steps of protein import take place.
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  • 39
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    Journal of bioenergetics and biomembranes 29 (1997), S. 35-43 
    ISSN: 1573-6881
    Keywords: Chaperonins ; heat-shock proteins ; mitochondria ; molecular chaperones ; protein folding ; protein import
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    Topics: Biology , Chemistry and Pharmacology , Physics
    Notes: Abstract Precursor proteins destined for the mitochondrial matrix traverse inner and outer organelle membranes in an extended conformation. Translocation events are therefore integrally coupled to the processes of protein unfolding in the cytosol and protein refolding in the matrix. To successfully import proteins from the cytoplasm into mitochondria, cells have recruited a variety of molecular chaperone systems and folding catalysts. Within the organelles, mitochondrial Hsp70 (mt-Hsp70) is a major player in this process and exerts multiple functions. First, mt-Hsp70 binds together with cohort proteins to incoming polypeptide chains, thus conferring unidirectionality on the translocation process, and then assists in their refolding. A subset of imported proteins requires additional assistance by chaperonins of the Hsp60/Hsp10 family. Protein folding occurs within the cavity of these cylindrical complexes. A productive interaction of precursor proteins with molecular chaperones in the matrix is not only crucial for correct refolding and assembly, but also for processing of presequences, intramitochondrial sorting, and degradation of proteins. This review focuses on the role of mt-Hsp70 and Hsp60/Hsp10 in protein folding in the mitochondrial matrix and discusses recent findings on their molecular mechanism of action.
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  • 40
    ISSN: 1573-6881
    Keywords: NADH: ubiquinone reductase ; ubiquinone ; proton pumping ; mitochondria
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    Topics: Biology , Chemistry and Pharmacology , Physics
    Notes: Abstract As part of the ongoing studies aimed at elucidating the mechanism of the energy conserving function of mitochondrial complex I, NADH: ubiquinone (Q) reductase, we have investigated how short-chain Q analogs activate the proton pumping function of this complex. Using a pH-sensitive fluorescent dye we have monitored both the extent and initial velocity of proton pumping of complex I in submitochondrial particles. The results are consistent with two sites of interaction of Q analogs with complex I, each having different proton pumping capacity. One is the physiological site which leads to a rapid proton pumping and a stoichiometric consumption of NADH associated with the reduction of the most hydrophobic Q analogs. Of these, heptyl-Q appears to be the most efficient substrate in the assay of proton pumping. Q analogs with a short-chain of less than six carbons interact with a second site which drives a slow proton pumping activity associated with NADH oxidation that is overstoichiometric to the reduced quinone acceptor. This activity is also nonphysiological, since hydrophilic Q analogs show little or no respiratory control ratio of their NADH:Q reductase activity, contrary to hydrophobic Q analogs.
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  • 41
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    Journal of bioenergetics and biomembranes 29 (1997), S. 97-102 
    ISSN: 1573-6881
    Keywords: Hexokinase ; binding to mitochondria ; mitochondria ; binding of hexokinase to ; Porin ; VDAC
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Physics
    Notes: Abstract Binding of the Type I isozyme of mammalian hexokinase to mitochondria is mediated by the porin present in the outer mitochondrial membrane. Type I hexokinase from rat brain is avidly bound by rat liver mitochondria while, under the same conditions, there is no significant binding to mitochondria from S. cerevisiae. Previously published work demonstrates the lack of significant interaction of yeast hexokinase with mitochondria from either liver or yeast. Thus, structural features required for the interaction of porin and hexokinase must have emerged during evolution of the mammalian forms of these proteins. If these structural features serve no functional role other than facilitating this interaction of hexokinase with mitochondria, it seems likely that they evolved in synchrony since operation of selective pressures on the hexokinase–mitochondrial interaction would require the simultaneous presence of hexokinase and porin capable of at least minimal interaction, and be responsive to changes in either partner that affected this interaction. Recent studies have indicated that a second type of binding site, which may or may not involve porin, is present on mammalian mitochondria. There are also reports of hexokinase binding to mitochondria in plant tissues, but the nature of the binding site remains undefined.
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  • 42
    ISSN: 1573-6881
    Keywords: ADP ; mitochondria ; free radical production ; brain ; heart ; exercise ; hypermetabolism
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    Topics: Biology , Chemistry and Pharmacology , Physics
    Notes: Abstract In agreement with classic studies, succinate-supplemented rat and pigeon heart and nonsynaptic brain mitochondrial free radical production is stopped by ADP additions causing the stimulation of respiration from State 4 to State 3. Nevertheless, with Complex I-linked substrates, mitochondria produce free radicals in State 3 at rates similar or somewhat higher than during resting respiration. The absence of sharp increases in free radical production during intense respiration is possible due to strong decreases of free radical leak in State 3. The results indicate that Complex I is the main mitochondrial free radical generator in State 3, adding to its already known important generation of active oxygen species in State 4. The observed rate of mitochondrial free radical production with Complex I-linked substrates in the active State 3 can help to explain two paradoxes: (a) the lack of massive muscle oxidative damage and shortening of life span due to exercise, in spite of up to 23-fold increases of oxygen consumption together with the very low levels of antioxidants present in heart, skeletal muscle, and brain; (b) the presence of some degree of oxidative stress during exercise and hyperactivity in spite of the stop of mitochondrial free radical production by ADP with succinate as substrate.
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  • 43
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    Journal of bioenergetics and biomembranes 29 (1997), S. 549-559 
    ISSN: 1573-6881
    Keywords: Luciferase ; localized probe ; heterogeneous coupled systems ; mitochondria ; hexokinase ; nucleotide concentration gradients ; cellular catalysis
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Physics
    Notes: Abstract The concentration of ATP generated by yeast mitochondria and consumed by yeast hexokinase was monitored using native firefly luciferase in solution, or recombinant luciferase localized at the surface of mitochondria. In the absence of hexokinase, both probes perform similarly in detecting exogenous or mitochondrially-generated ATP. The steady-state concentrations of ATP can be reduced in a dose-dependent manner by hexokinase. With hexokinase added in large excess, the localized probe reports substantial ATP concentrations while none is detectable by soluble luciferase. Thus, ATP accumulates near the membrane where it appears, relatively to solution, and vice versa for ADP. The extent of nucleotide gradients is shown to be correlated with the specific activity of oxidative phosphorylation and with the viscosity of the medium, but independent of the concentration of the organelles. A simple model involving diffusional restrictions is presented to describe this behavior. The metabolic and evolutionary implications of cellular catalysis limitation by physical processes are discussed.
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    Journal of bioenergetics and biomembranes 29 (1997), S. 233-239 
    ISSN: 1573-6881
    Keywords: ATP synthase subunit 8 ; genes ; mammals ; mitochondria ; sea urchins ; sequences
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Physics
    Notes: Abstract Peculiar evolutionary properties of the subunit 8 of mitochondrial ATP synthase (ATPase8) are revealed by comparative analyses carried out between both closely and distantly related species of echinoderms. The analysis of nucleotide substitution in the three echinoids demonstrated a relaxation of amino acid functional constraints. The deduced protein sequences display a well conserved domain at the N-terminus, while the central part is very variable. At the C-terminus, the broad distribution of positively charged amino acids, which is typical of other organisms, is not conserved in the two different echinoderm classes of the sea urchins and of the sea stars. Instead, a motif of three amino acids, so far not described elsewhere, is conserved in sea urchins and is found to be very similar to the motif present in the sea stars. Our results indicate that the N-terminal region seems to follow the same evolutionary pattern in different organisms, while the maintenance of the C-terminal part in a phylum-specific manner may reflect the co-evolution of mitochondrial and nuclear genes.
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    Journal of bioenergetics and biomembranes 29 (1997), S. 303-313 
    ISSN: 1573-6881
    Keywords: Hepatic preneoplasia ; glycogenotic foci ; amphophilic foci ; mitochondria ; peroxisomes ; hepatocellular neoplasms
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Physics
    Notes: Abstract Biochemical and molecular biological approaches in situ have provided compelling evidence for early bioenergetic changes in hepatocarcinogenesis. Hepatocellular neoplasms regularly develop from preneoplastic foci of altered hepatocytes, irrespective of whether they are caused by chemicals, radiation, viruses, or transgenic oncogenes. Two striking early metabolic aberrations were discovered: (1) a focal excessive storage of glycogen (glycogenosis) leading via various intermediate stages to neoplasms, the malignant phenotype of which is poor in glycogen but rich in ribosomes (basophilic), and (2) an accumulation of mitochondria in so-called oncocytes and amphophilic cells, giving rise to well-differentiated neoplasms. The metabolic pattern of human and experimentally induced focal hepatic glycogenosis mimics the phenotype of hepatocytes exposed to insulin. The conversion of the highly differentiated glycogenotic hepatocytes to the poorly differentiated cancer cells is usually associated with a reduction in gluconeogenesis, an activation of the pentose phosphate pathway and glycolysis, and an ever increasing cell proliferation. The metabolic pattern of preneoplastic amphophilic cell populations has only been studied to a limited extent. The few available data suggest that thyromimetic effects of peroxisomal proliferators and hepadnaviral infection may be responsible for the emergence of the amphophilic cell lineage of hepatocarcinogenesis. The actions of both insulin and thyroid hormone are mediated by intracellular signal transduction. It is, thus, conceivable that the early changes in energy metabolism during hepatocarcinogenesis are the consequence of alterations in the complex network of signal transduction pathways, which may be caused by genetic as well as epigenetic primary lesions, and elicit adaptive metabolic changes eventually resulting in the malignant neoplastic phenotype.
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    Journal of bioenergetics and biomembranes 29 (1997), S. 151-163 
    ISSN: 1573-6881
    Keywords: cytochrome c oxidase ; respiratory chain ; mitochondria ; assembly ; enzyme deficiency ; Leigh's syndrome ; mitochondrial myopathy (Saccharomyces cerevisiae, human)
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Physics
    Notes: Abstract As the terminal component of the mitochondrial respiratory chain, cytochrome c oxidase plays a vital role in cellular energy transformation. Human cytochrome c oxidase is composed of 13 subunits. The three major subunits form the catalytic core and are encoded by mitochondrial DNA (mtDNA). The remaining subunits are nuclear-encoded. The primary sequence is known for all human subunits and the crystal structure of bovine heart cytochrome c oxidase has recently been reported. However, despite this wealth of structural information, the role of the nuclear-encoded subunits is still poorly understood. Yeast cytochrome c oxidase is a close model of its human counterpart and provides a means of studying the effects of mutations on the assembly, structure, stability and function of the enzyme complex. Defects in cytochrome c oxidase function are found in a clinically heterogeneous group of disorders. The molecular defects that underlie these diseases may arise from mutations of either the mitochondrial or the nuclear genomes or both. A significant number of cytochrome c oxidase deficiencies, often associated with other respiratory chain enzyme defects, are attributed to mutations of mtDNA. Mutations of mtDNA appear, nonetheless, uncommon in early childhood. Pedigree analysis and cell fusion experiments have demonstrated a nuclear involvement in some infantile cases but a specific nuclear genomic lesion has not yet been reported. Detailed analyses of the many steps involved in the biogenesis of cytochrome c oxidase, often pioneered in yeast, offer several starting points for further molecular characterizations of cytochrome c oxidase deficiencies observed in clinical practice.
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    Journal of bioenergetics and biomembranes 29 (1997), S. 331-338 
    ISSN: 1573-6881
    Keywords: Cancer ; proliferation ; Crabtree effect ; insulin action ; compartmentation ; aerobic glycolysis ; hexokinase ; mitochondria ; porin ; protein synthesis ; TCA cycle
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Chemistry and Pharmacology , Physics
    Notes: Abstract Current thought is that proliferating cells undergo a shift from oxidative to glycolytic metabolism, where the energy requirements of the rapidly dividing cell are provided by ATP from glycolysis. Drawing on the hexokinase–mitochondrial acceptor theory of insulin action, this article presents evidence suggesting that the increased binding of hexokinase to porin on mitochondria of cancer cells not only accelerates glycolysis by providing hexokinase with better access to ATP, but also stimulates the TCA cycle by providing the mitochondrion with ADP that acts as an acceptor for phosphoryl groups. Furthermore, this acceleration of the TCA cycle stimulates protein synthesis via two mechanisms: first, by increasing ATP production, and second, by provision of certain amino acids required for protein synthesis, since the amino acids glutamate, alanine, and aspartate are either reduction products or partially oxidized products of the intermediates of glycolysis and the TCA cycle. The utilization of oxygen in the course of the TCA cycle turnover is relatively diminished even though TCA cycle intermediates are being consumed. With partial oxidation of TCA cycle intermediates into amino acids, there is necessarily a reduction in formation of CO2 from pyruvate, seen as a relative diminution in utilization of oxygen in relation to carbon utilization. This has been assumed to be an inhibition of oxygen uptake and therefore a diminution of TCA cycle activity. Therefore a switch from oxidative metabolism to glycolytic metabolism has been assumed (the Crabtree effect). By stimulating both ATP production and protein synthesis for the rapidly dividing cell, the binding of hexokinase to mitochondrial porin lies at the core of proliferative energy metabolism. This article further reviews literature on the binding of the isozymes of hexokinase to porin, and on the evolution of insulin, proposing that intracellular insulin-like proteins directly bind hexokinase to mitochondrial porin.
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    Colloid & polymer science 106 (1997), S. 192-197 
    ISSN: 1435-1536
    Keywords: ADP/ATP carrier ; loopstructure ; substrate binding site ; SH-reagent ; mitochondria
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology , Mechanical Engineering, Materials Science, Production Engineering, Mining and Metallurgy, Traffic Engineering, Precision Mechanics
    Notes: Abstract The transport mechanism of the bovine heart mitochondrial ADP/ATP carrier was studied using submitochondrial particles. The modifications of the cysteine residues of the carrier by the SH-reagents eosin-5-maleimide (EMA) andN-ethylmaleimide (NEM), and disulfide bond formation between these cysteine residues catalyzed by copper-o-phenanthroline (Cu(OP)2) under various conditions were studied. In particular, the effects of the transport inhibitors carboxyatractyloside (CATR) and bongkrekic acid (BKA), and fluorescein derivatives were examined. From the results, the topology of the carrier in the membrane, dynamic translocations of the loops of the carrier, and the structure of the primary binding site of the transport substrates ADP and ATP were deduced. The loops are concluded to act as both gates in the transport and binding sites for the substrates. Based on the results, a cooperative swinging-loop model is postulated as the transport mechanism of the ADP/ATP carrier.
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    Plant molecular biology 34 (1997), S. 823-829 
    ISSN: 1573-5028
    Keywords: sorghum ; mitochondria ; transcription ; trnfM ; atp9
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Sorghum mitochondrial atp9 is polymorphic among male-sterile cytoplasms, but each cytoplasm is characterized by a major 650 nt transcript, regardless of fertility status. The gene is positioned 323 bp 3′ to trnfM. Primer extension revealed multiple atp9 5′ transcript termini, distributed from +1 to +28 3′ to trnfM; the termini could be labeled with polynucleotide kinase, suggesting that they result from the maturation of trnfM. Guanylyltransferase experiments, however, showed that four of the termini were capable. The juxtaposition of a putative promoter 3′ to trnfM results in a unique atp9 transcript population consisting of primary and processed transcripts.
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    World journal of microbiology and biotechnology 14 (1997), S. 191-197 
    ISSN: 1573-0972
    Keywords: Candida boidinii ; catalase ; electron microscopy ; methanol-utilizing yeasts ; mitochondria
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract Catalase activity of the methanol-assimilating yeast Candida boidinii M-363 was determined cytochemically and biochemically. Electron microscopic investigations on ultrathin sections were made on cells from 16, 24, and 48h batch cultures in nutrient medium with methanol (or glucose as a control) as the sole source of carbon and energy. The electron-dense oxidation product of 3,3′-diaminobenzidine was found predominantly in the mitochondrial cristae and membranes. The mitochondria were increased in number, enlarged, sometimes aggregated, with variable form and size and they characteristically developed when the strain was grown on methanol. The significant development of these organelles and their intensive DAB staining correlated with the considerable increase in catalase activity. Biochemically, catalase in the cell-free extract was determined to be maximal along the exponential growth phase of the strain during its incubation on methanol. Enzyme analysis of the heavy mitochondrial fraction showed that it possessed catalase activity but not peroxidase activity. The results showed that not only peroxisomes but also mitochondria may be structurally and functionally responsible for the high catalase activity of some methanol-assimilating yeasts. What is more, the contribution of the mitochondria to the utilization of methanol may be significant.
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    Bulletin of experimental biology and medicine 124 (1997), S. 658-660 
    ISSN: 1573-8221
    Keywords: phospholipids ; mitochondria ; liver ; hemorrhagic shock
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Heparin used as an anticoagulant in modeled hemorrhagic shock decreases the phosphatidylcholine and increases the phosphatidylethanolamine contents in the mitochondria. Accumulation of lysophosphatidylcholine in whole mitochondria and their inner membrane is observed in hemorrhagic shock. At the same time, hemorrhagic shock decreases phosphatidylcholine content in the inner and outer mitochondrial membranes and increases phosphatidylethanolamine content in the outer membranes. Modification of phospholipid composition of mitochondrial membranes is a mechanism responsible for impaired energy production in liver mitochondria in hemorrhagic shock.
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    Biologia plantarum 39 (1997), S. 607-614 
    ISSN: 1573-8264
    Keywords: chloroplasts ; fern ; mitochondria ; photosynthesis ; respiration ; salt stress ; ultrastructure
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Gametophytes cultured in solutions containing 0.0 to 0.7 % NaCl exhibited no change in ultra structural organization of chloroplasts. In 1.0% NaCl-grown gametophytes, there were thinner granal stacks, relatively larger spaces between granal thylakoidal membranes and larger plastoglobuli in the chloroplasts. These changes were accompanied by a decrease in photosynthesis. Cup shape, horseshoe shape, ring shape, and amoeboid mitochondria were observed in gametophytes grown in 0.0 to 0.7% NaCl. Only round mitochondria were observed in the gametophytes grown in 1.0 % NaCl. Mitochondria seemed to be more resistant to salt stress compared to chloroplasts. There was no direct relationship between changes in respiration rate and changes in mitochondrial shape among gametophytes grown in different NaCl concentrations.
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    Transgenic research 6 (1997), S. 379-383 
    ISSN: 1573-9368
    Keywords: genetic engineering ; heteroplasmy ; mouse ; mitochondria ; mitochondria transfer
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract A method for mitochondria isolation and interspecific transfer of mitochondria was developed in mice. Mitochondria were isolated from Mus spretus liver samples for microinjection into fertilized ova obtained from superovulated M. musculus domesticus females. Electron microscopic observations of mitochondria preparations used for microinjection demonstrated intact mitochondrial vesicles with little microsomal contamination. Species-specific nested PCR primers complementary to sequence differences in the mitochondrial DNA D-loop region revealed high rates of successful transfer of foreign mitochondria after isolation and injection into zygotes cultured through the blastocyst stage of embryonic development. Of 217 zygotes, 67 survived mitochondria injection and 23 out of 37 zygotes developed were at the blastocyst-stage of embryonic development after 4.5 days of in vitro culture. All 23 of these blastocysts contained detectable levels of foreign mitochondria. These results represent an initial step in developing a model system to study mitochondrial dynamics and development of therapeutic strategies for human metabolic diseases affected by aberrations in mitochondrial function or mutation
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