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Biomedical subjects

F M Goñi

Publications and source records attributed to F M Goñi.

At least 127 records · Page 7Linked to original sources
PubMed1985

Phospholipid oxidation catalyzed by cytochrome c in liposomes.

Oxidation of liposome phospholipids has been studied in the presence of cytochrome c. Sonicated vesicles of soya bean or egg-yolk lipids, or purified phospholipid preparations, were treated with oxidized cytochrome c at a 10:4 lipid/protein ratio (w/w). Lipid peroxidation was examined by oxygen polarography, gas-liquid chromatography (GLC) and the thiobarbituric acid test. Oxidized, but not reduced, cytochrome effectively catalyzes lipid oxidation under these conditions. Oxygen consumption and disappearance of unsaturated fatty acids follow closely similar patterns, the O2 consumption rate showing a maximum (1.53 mol O2/min per mol heme) shortly before fatty acid loss reaches its peak. GLC and O2 consumption data suggest that monohydroperoxides are the most abundant oxidized species in the system. The thiobarbituric acid reaction, however, appears only to be of qualitative value in peroxidation studies. In order to test the mechanism through which oxidation occurs in our system, the effect of liposome composition and the presence of antioxidants was tested, both on cytochrome c binding to bilayers and on O2 consumption. Oxidized and reduced cytochrome c bind the lipid bilayers with similar affinity, but only the oxidized form is active in autoxidation. Antioxidants do not modify either cytochrome c binding to sonicated liposomes. Lipid composition does influence considerably cytochrome binding, and O2 consumption is correspondingly altered. Studies with various antioxidants and inhibitors suggest that both free radicals and singlet oxygen may be involved in the process under study.

Animals↗
PubMed1985

Protein conformational transitions in sarcoplasmic reticulum membranes.

Fourier-transform infrared spectroscopic studies of sarcoplasmic reticulum proteins, in H2O and D2O, suggest that 10 mM ATP induces a conformational change in those proteins, increasing their contents in alpha-helical and beta-antiparallel structures. Ca2+ on the contrary, is seen to reduce the proportion of alpha-helix and increase the contribution of random coil.

Adenosine Triphosphate↗
PubMed1985

Laser light-scattering characterization of mitochondrial complex III-Triton X-100-phospholipid mixed micelles.

Bovine-heart mitochondrial complex III was purified in the presence of Triton X-100, and the size and shape of the resulting protein-surfactant-phospholipid mixed micelles were investigated by laser light-scattering. The protein appears to be present in the form of a dimer, irrespective of temperature (between 25 and 40 degrees C) and protein concentration (between 0.5 and 5 mg/ml). The molecular weight of the micelle increases with temperature from 600 000 (25 degrees C) to 692 000 (40 degrees C). The variation of the solvent second virial coefficient in this temperature range suggests that, with increasing temperature, some of the free surfactant molecules become integrated in the mixed micelles. The average quadratic radius of gyration of these is of 42 +/- 5 nm, corresponding in our case to an ellipsoidal shape.

Animals↗
PubMed1985

The effect of bilayer order and fluidity on detergent-induced liposome fusion.

Surfactants induce fusion (or increase in size) of sonicated liposomes. This phenomenon is enhanced by cholesterol and inhibited by the intrinsic polypeptide gramicidin A. By comparison with previous physical studies we conclude that liposome 'fusion' is facilitated when both fluidity and static order of the bilayer are high.

Cholesterol↗
PubMed1985

Colorimetric assay of gramicidin A in the presence of surfactants and phospholipids.

Gramicidin A, a hydrophobic polypeptide containing 4 tryptophan residues/molecule, may be determined quantitatively after reaction with 4-(dimethylamino)benzaldehyde, a method previously proposed for tryptophan analysis. The assay may be carried out even in the presence of various surfactants and phospholipids at high concentrations.

Benzaldehydes↗
PubMed1984

Infrared spectroscopy of phosphatidylcholines in aqueous suspension. A study of the phosphate group vibrations.

The 1000-1300 cm-1 region of the infrared spectrum of dipalmitoylphosphatidylcholine (DPPC) and other phosphate-containing molecules has been studied by the Fourier-transform technique. Three absorption bands have been assigned to various vibrational modes of the DPPC phosphate group, with maximum wavenumbers at 1060, 1086 and 1222 cm-1. These values are the same above and below Tc of the phospholipid. Dehydration produces band-shifts toward higher wavenumbers .

Fourier Analysis↗
PubMed1984

Physiological state of submitochondrial particles and their susceptibility to Triton X-100.

The solubilizing effect of Triton X-100 on beef heart submitochondrial particles (ETPH) has been studied under various physiological conditions. Coupled, uncoupled and azide-inhibited ETPH particles have been studied. Quantitative and qualitative differences are found in the proteins solubilized by the detergent from ETPH particles under the various conditions tested.

Animals↗
PubMed1984

Effect of the nonionic detergent Triton X-100 on mitochondrial succinate-oxidizing enzymes.

Specific activities of succinate:coenzyme Q reductase, ubiquinone:cytochrome c reductase, cytochrome oxidase, succinate:cytochrome c reductase, succinate oxidase, and ubiquinol oxidase have been measured in rat liver mitochondria in the presence of Triton X-100. The last three activities are much more sensitive to Triton X-100 than the first ones; the data suggest that the electron transport chain components cannot react with each other in the presence of the detergent. At least in the case of succinate:cytochrome c reductase, reconstitution of the detergent-treated membranes with externally added phospholipids reverses the inhibition produced by Triton X-100. These results support the idea that the respiratory chain components diffuse at random in the plane of the inner mitochondrial membrane; the main effect of the detergent would be to impair lateral diffusion by decreasing the area of lipid bilayer. When detergent-treated mitochondrial suspensions are centrifuged in order to separate the solubilized from the particulate material, only the first three enzyme activities mentioned above are found in the supernatants. After centrifugation, a latent ubiquinol:cytochrome c oxidase activity becomes apparent, whereas the same centrifugation process produces inhibition of cytochrome c oxidase in the presence of certain Triton X-100 concentrations. These effects could be due either to a selective solubilization of regulatory or catalytic subunits or to a conformational change of the enzyme-detergent complex.

Animals↗
PubMed1984

Lipid composition and protoplast-forming capacity of Streptomyces antibioticus.

The lipid and fatty acid composition of a strain of Streptomyces antibioticus has been studied as a function of culture age and glycine concentration in the growth medium. Under all conditions, the main polar lipids were phosphatidylethanolamine, cardiolipin and phosphomannoinositides in order of decreasing abundance; no ornithinolipids were detected. Acylglucoses and menaquinones were found among the nonpolar lipids. The main fatty acids present were anteiso 15:0 and anteiso 17:0. The lipid composition of the cells varied with the age of the culture, but no uniform pattern of variation was found in the cultures grown on different amounts of glycine. Among the cells harvested at the end of the exponential phase of growth, those grown on 2% glycine give the highest yield of protoplast formation. These cells were found to contain low amounts of nonpolar lipids and of phosphatidylethanolamine, and high proportions of anteiso fatty acids. We propose that the membrane bilayer of these cells, because of its peculiar lipid composition, is particularly stable and fluid. As a consequence, the yield and stability of derived protoplasts should be increased.

Fatty Acids↗
PubMed1983

Membrane-surfactant interactions. The effect of Triton X-100 on sarcoplasmic reticulum vesicles.

The effect of Triton X-100 on purified sarcoplasmic reticulum vesicles has been studied by means of chemical, ultrastructural and enzymic techniques. At low detergent/membrane ratios (about 1 Triton X-100 per 60 phospholipid molecules) the only effect observed is an increase in vesicle permeability. Higher surfactant concentrations, up to a 1:1 detergent/phospholipid ratio, produce a large enhancement of ATPase activity. Membrane solubilization occurs as a critical phenomenon when the surfactant/phospholipid molar ratio reaches a value around 1.5:1, corresponding to 2 mumol Triton X-100/mg protein. At this point, the suspension turbidity drops, virtually all the protein and phospholipid is solubilized and every organized structure disappears. Simultaneously, a dramatic increase in the specific activity of the solubilized ATPase is observed. The sudden solubilization of almost all the bilayer components at a given detergent concentration is attributed to the relative simplicity of this membrane system. Solubilization takes place at the same surfactant/membrane ratio, at least between 0.5 and 4 mg membrane protein/ml. The non-solubilized residue seems to consist mainly of delipidized aggregated forms of ATPase.

Animals↗
PubMed1983

The impairment of mitochondrial function by Triton X-100. A study of mitochondrial respiration and energy-dependent swelling.

Coupled and uncoupled respiration, and energy-dependent phosphate swelling have been studied in rat liver mitochondria in the presence of various concentration of Triton X-100. Detergent concentrations up to 10(-5) M do not affect any of the processes under study. At 10(-5) M, Triton X-100 produces a slight decrease of coupled respiration and a considerable inhibition of mersalyl-induced shrinking in swollen mitochondria. Increasing the surfactant concentration to 10(-4), coupled as well as uncoupled O2 consumption is decreased, succinate-dependent phosphate swelling is inhibited and an energy-dependent phosphate swelling in the absence of valinomycin is observed. At 2 X 10(-4) M. Triton X-100, ATP- dependent phosphate swelling is abolished, and passive swelling may be induced by various ions. Higher detergent concentrations do not allow observation of any of these events. On the basis of these results, a model of membrane-detergent interaction is proposed.

Animals↗
PubMed1982

A comparative study of the effect of various detergents on the structure and function of sarcoplasmic reticulum vesicles.

The effects produced by the detergents Triton X-100, sodium dodecylsulphate and sodium cholate on sarcoplasmic reticulum vesicles have been comparatively studied. In all cases, maximal effects are found 5 min after detergent addition. Triton X-100 and SDS are approximately ten times more effective than cholate in protein and phospholipid solubilization. Both Triton X-100 and SDS maintain Ca++ accumulation in SR vesicles at detergent concentrations below 10(-3) M; higher concentrations cause a strong inhibition. On the other hand, cholate produces a gradual inhibition of Ca++ accumulation in the concentration range between 10(-4) M and 2.5 X 10(-2) M. Triton X-100 and SDS produce a gradual solubilization of the specific Ca++-ATPase activity up to a 10(-3) M detergent concentration, above which a strong inactivation occurs, while the enzyme solubilization increases with the presence of cholate in the whole concentration range under study. The different behaviour of sodium cholate, when compared to SDS or Triton X-100, is discussed in relation to the surfactant molecular structures. The possibility of membrane lysis and reassembly in the presence of some detergents is also considered.

Animals↗
PubMed1982

Increase in size of sonicated phospholipid vesicles in the presence of detergents.

Triton X-100, sodium dodecylsulphate, sodium cholate, and beta-octylglucoside increase the size of sonicated, but not of unsonicated, phospholipid vesicles above the Tc gel to liquid-crystalline transition temperature. Lysophosphatidylcholine or glyceryl monooleate do not increase liposome size under these circumstances. The observed phenomenon of vesicle growth is virtually unaffected by phospholipid composition, surface potential, calcium ions, EDTA, or albumin. The presence of cholesterol makes the vesicles more susceptible to detergent-induced growth. The presence of detergents, even at concentrations much lower than required to observe any increase in vesicle size, favors the liberation of vesicle contents. These results may have implications concerning membrane fusion as well as the reconstitution of membrane proteins in the presence of detergents.

Calcium↗
PubMed1982

Fractionation of rat liver mitochondrial components after short treatments with Triton X-100.

1. When rat liver mitochondria are treated with the non-ionic detergent Triton X-100, the solubilizing effects of the latter are already maximal after a few seconds. 2. We have developed a filtration technique that allows a fast separation of the solubilized and non-solubilized fractions. By means of this technique, we have been able to show differences in the solubilizing effect of Triton X-100 according to the physiological state of mitochondria. 3. In addition, the filtrate may be subjected to further fractionation by ultracentrifugation, by which two filtrate subfractions, supernatant and pellet, may be obtained. 4. Filtrates, supernatants and pellets differ from each other, and from intact mitochondria, in phospholipid and polypeptide composition, and possess a characteristic distribution of enzyme activities; they are also different from the ultrastructural point of view. 5. It is concluded that the combined techniques of short detergent treatments and fast filtration are useful in structural and functional studies of mitochondria and in the selective solubilization of mitochondrial components.

Animals↗