Properties of mitochondria from cells of the "fermentative" variant of Endomyces magnusii]

The properties of mitochondria from the cells of the "fermentative" variant of End. magnusii were studied. The induced fermentative transformation was brought about by a non-balanced vitamin cultivation. It was shown that the "fermentative" variant of End. magnusii represents an interesting model, in which the energy required for the cell functioning is provided for by a high fermentative activity and a normally functioning respiratory chain. The "fermentative" variant mitochondria were tightly coupled and possessed theoretical efficiency during oxidation of NAD-dependent substrates, which suggested the existence of all the three sites of energy coupling and phosphorylation at the substrate level. A specificity of energy regulation of the End. magnusii "fermentative" variant mitochondria, e. g. tight coupling during oxidation of succinate and lack of tight coupling during oxidation of exogenous NADH, is discussed. The tight coupling during succinate oxidation is confirmed by the observation of reverse electron transfer. Thus, the energy-dependent reduction of NAD during succinate oxidation has been firstly demonstrated for the mitochondria of yeast grown on a fermentable substrate.     

Methane oxidation potential in the water column of two diverse coastal marine sites

Methane oxidation in the water column was investigated at two nearshore marine environments with relatively high concentrations of dissolved methane. In the northern Gulf of Mexico, high methane oxidation rates were observed at the pycnocline, with the highest oxidation rate corresponding to the most negative bacterial ¹³C values. These low isotopic values occurred during the winter when overall bacterial productivity was low, suggesting that at this time of the year, methanotrophs in the Gulf could make up a significant portion of the overall bacterial assemblage. Although methane oxidation also occurred during more productive times (i.e., summer), the isotopic signal of methane oxidation was not observed in the bacterial biomass because of the higher overall bacterial productivity. The other site, Cape Lookout Bight, NC, is a small marine embayment where methane is produced in the organic-rich sediments. No measurable rates of methane oxidation in the water column occurred, and no anomalously low ¹³C values of the bacterioplankton were measured. In both environments, methane production and oxidation appear to be spatially coupled, occurring at/near the pycnocline in the northern Gulf of Mexico and at the sediment-water interface at Cape Lookout Bight, NC.     

Role of -NADH for microsomal ethanol oxidation

α-NADH¹ was found to serve as electron donor for microsomal ethanol oxidation of rat liver. Almost no ethanol oxidation was observed with β-NADH. The α-NADH-dependent ethanol oxidation was almost completely inhibited by 0.1 mM cyanide or azide and strongly abolished in the presence of formate. α-NADH-dependent ethanol oxidation was increased by 1 mM SKF-525A, an inhibitor of microsomal mixed-function oxidase, to about 200%.These results suggested that hydrogen peroxide generated from α-NADH and molecular oxygen in microsomes might be a prerequisite step in the over-all reaction, eventually leading to the peroxidatic ethanol oxidation by catalase to acetaldehyde.     

Gender-based differences in substrate use during exercise at a self-selected pace

The aim of this study was to investigate gender-based differences in substrate use during exercise at a self-selected pace. Seventeen men and 17 women performed a maximal exercise test and a 20-minute bout of self-paced treadmill walking to determine carbohydrate and fat oxidation rates. Gas exchange measurements were performed throughout the tests, and stoichiometric equations were used to calculate substrate oxidation rates. For each individual, a best-fit polynomial curve was constructed using fat oxidation rate (g·min(-1)) vs. exercise intensity (percentage of maximal oxygen uptake, % VO(2)max). Each individual curve was used to obtain the following variables: maximal fat oxidation (MFO), the peak rate of fat oxidation measured over the entire range of exercise intensities; fat(max), the exercise intensity at which the MFO was observed; and fat(max) zone, range of exercise intensities with fat oxidation rates within 10% of fat oxidation rates at fat(max). Although the MFO was similar between genders, fat(max) was lower in men than in women. Similarly, the "low" and "high" borders of the fat(max) zone were lower in men than in women. During exercise at a self-selected pace, carbohydrate oxidation rates were greater in men than in women, despite no gender-based differences in fat oxidation rates. However, fat oxidation contribution to total energy expenditure (EE) was greater in women than in men, despite no gender-based differences in the exercise intensity. In conclusion, although both genders self-selected a similar exercise intensity, the contribution of fat oxidation to EE is greater in women than in men. Interestingly, both genders self-selected an exercise intensity that falls within the fat(max) zone.     

Oxidation of sulfur compounds and electron transport in Thiobacillus denitrificans

Summary Intact cells of Thiobacillus denitrificans catalyzed the oxidation of thiosulfate, sulfide and sulfite with nitrate or oxygen as the terminal acceptor. The anaerobic oxidation of thiosulfate, sulfide and sulfite was sensitive to the inhibitors of the flavoprotein system. Under aerobic conditions the oxidation of sulfide and sulfite was sensitive to these inhibitors but the thiosulfate oxidation was unaffected. Cyanide and azide inhibited the aerobic and anaerobic respiration when thiosulfate, sulfide or sulfite served as electron donors. The oxidation of thiosulfate by cell-free preparations was mediated by cytochromes of c, a and o-types. The cell-free extracts also catalyzed the oxidation of NADH and succinate, involving flavoproteins and b, c, a and o-type cytochromes. In addition, a cytochrome oxidase sensitive to cyanide and azide was also present.Non-Standard Abbreviations TTFA Thenoyltrifluoroacetone - HQNO 2-heptyl-4-hydroxyquonoline N-oxide Aspirant van het Nationaal Fonds voor Wetenschappelijk Onderzoek (Belgian National Science Foundation).     

Some quantitative histochemical studies of the periodic acid oxidation of glycogen in situ

Synposis The periodic acid oxidation of the glycogens in sections of ox liver fixed in Carnoy's fluid has been studied quantitatively. It was found that in 0.02 M acetate buffer at pH 5.0 the oxidation was rapid at first but levelled off after I hr. Even after prolonged oxidation (12 days), not more than 23–24% of the total available glycogen was oxidized. However, in the presence of electrolytes (e.g. 0.2.m sodium chloride) the oxidation was much greater. After 2.5 hr, for example, 40% of the available glycogen was oxidized. There was little difference in the velocity of oxidation in sections mounted on glass-slides and free-floating ones, or in free-floating sections of different thicknesses.Mounted and unmounted sections consumed on average 11–17 times more periodate than could be accounted for by the oxidation of their glycogen content.The results are interpreted in terms of a complex formed between periodate ions and the cuter glucosyl residues of the glycogen aggregate. The negative charge on the complex, it is suggested, prevents free periodate ions approaching and oxidizing the inner glycosyl residues.     

Catabolic enzyme activities in the pectoralis muscle of premigratory and migratory juvenile Reed Warblers Acrocephalus scirpaceus (Herm.)

Summary Differences in catabolic capacity in the pectoralis major muscle of premigratory and migratory Reed Warblers were examined. The oxidative capacity was greater in the migratory birds, probably reflecting a training effect caused by the increased locomotion activity just prior to migration.Fatty acid oxidation was already high in the premigratory birds, possibly reflecting a saving of carbohydrate for anabolic purposes (feather growth) during the moult. Glycolytic capacity was slightly increased in the migratory birds, suggesting an increased carbohydrate oxidation during migration in spite of the great contribution of fatty acid oxidation.     

Redox chemistry in laccase-catalyzed oxidation of N-hydroxy compounds

1-Hydroxybenzotriazole, violuric acid, and N-hydroxyacetanilide are three N-OH compounds capable of mediating a range of laccase-catalyzed biotransformations, such as paper pulp delignification and degradation of polycyclic hydrocarbons. The mechanism of their enzymatic oxidation was studied with seven fungal laccases. The oxidation had a bell-shaped pH-activity profile with an optimal pH ranging from 4 to 7. The oxidation rate was found to be dependent on the redox potential difference between the N-OH substrate and laccase. A laccase with a higher redox potential or an N-OH compound with a lower redox potential tended to have a higher oxidation rate. Similar to the enzymatic oxidation of phenols, phenoxazines, phenothiazines, and other redox-active compounds, an "outer-sphere" type of single-electron transfer from the substrate to laccase and proton release are speculated to be involved in the rate-limiting step for N-OH oxidation.     

Utilization of sulfide and elemental sulfur by Ectothiorhodospira halochloris

Ectothiorhodospira halochloris grows photoheterotrophically with a variety of sulfur sources. During sulfide oxidation to elemental sulfur considerable amounts of polysulfides may be accumulated transiently. When grown on elemental sulfur no sulfate was produced by oxidation, but sulfide and polysulfide were formed by reduction. Only one soluble cytochrome c-551 was isolated and purified. It was a small acidic hemeprotein with a molecular weight of 6,300, an isoelectric point of 3.1 and a redox potential of-11 mV at pH 7.0. It showed three absorption maxima in the reduced state (=551 nm; =523 nm; =417 nm). The addition of various c-type cytochromes to a suspension of spheroplasts stimulated the velocity of sulfide oxidation. This stimulation was best with the small acidic cytochromes from E. halochloris or Ectothiorhodospira abdelmalekii. Sulfide oxidation was stopped by several uncoupling agents, ionophores and electron transport inhibitors. Antimycin A, rotenone and cyanide had no effect on sulfide oxidation.Dedicated to Prof. Dr. H. G. Schlegel on the occasion of his 60th birthday     

Oxidation of polycyclic aromatic hydrocarbons by laccase of Coriolus hirsutus

The oxidation of five polycyclic aromatic hydrocarbons; anthracene, benzo()pyrene, fluoranthene, phenanthrene and pyrene was catalyzed by laccase from Coriolus hirsutus in the presence of the redox mediators, 2,2-azinobis(3-ethylbenzthiazoline-6-sulfonate) (ABTS) and 1-hydroxybenzotriazole (HBT). In the ABTS-mediated system, benzo()pyrene was the most rapidly oxidized substrate, with anthracene being the most rapidly oxidized in the HBT-mediated system. There was no clear relationship between the ionization potential and the oxidation of the substrates. ABTS increased the oxidation of benzo()pyrene more than HBT but the oxidation of the other PAHs tested were the opposite. The mediators used in conjunction increased the oxidation of benzo()pyrene compared to using the mediators alone.     

Release of fossil methane from mineral soil particles, and its implication for estimation of methane oxidation in a mineral subsoil

In a preliminary experiment we found that methane evolved from a sandy subsoil during aerobic incubation of shaken soil slurries. In the study presented here the methane was found to be released from the sand particles by mechanical weathering, caused by the grinding effect of the shaking. Large amounts of gas (about 0.5 ml gas g^–1 soil) were extracted by intense grinding of the soil in gas tight serum vials. Methane was the main hydrocarbon in the emitted gas, but also a considerable amount of ethane was present, as well as minor amounts of heavier hydrocarbons (up to C₆). The ¹³C-values of the emitted methane and ethane were –33 and –29 , respectively. Together these results demonstrate a thermogenic origin of the gas. This paper also reports the results of an incubation experiment where possible methane oxidation was looked for. If a possible release of methane is not accounted for, methane oxidation may be overlooked, as illustrated in this paper. Methane consumption was detected only in soil from 40 cm, in contrast to soil sampled at 100 cm and deeper where a slight production was measured. When methane oxidation was inhibited by dimethyl-ether, a significant release of methane was seen. The release was probably caused by chemical weathering. When this methane release was taken into account, methane oxidation was found to be present at all measured depths (40 to 200 cm). Fertilization with urea inhibited the methane oxidation at 40 cm but not at deeper layers. It is hypothesized that ammonia oxidizing bacteria were the main methane oxidizers in this mineral subsoil (deeper than 1 m), and that oxidation of methane might be a survival mechanism for ammonia oxidizers in ammonia limited environments.     

Study of amino acid formation during palmitate oxidation in rat brain mitochondria

The interrelation of palmitate oxidation with amino acid formation in rat brain mitochondria has been investigated in purified mitochondria of nonsynaptic origin by measuring the formation of aspartate, -ketoglutarate, and glutamate during palmitate oxidation, and also by assaying¹⁴C-products of [1-¹⁴C]palmitate oxidation. Oxidation of palmitate (or [1-¹⁴C]palmitate) resulted in the formation of aspartate (or¹⁴C-aspartate), and the oxidation was inhibited by aminooxyacetate (an inhibitor of transaminase), Palmitate oxidation also resulted in -ketoglutarate formation, which was sensitive to the effect of aminooxyacetate. Addition of NH₄Cl was found to increase¹⁴C-products and formation of -ketoglutarate, whereas glutamate formation was not increased unless the rate of palmitate oxidation was reduced by 50% by aminooxyacetate or -ketoglutarate was added exogenously. Exogenous -ketoglutarate was found to decrease¹⁴C-products, but not aspartate formation. These results indicated that palmitate oxidation was closely related to aspartate formation via aspartate aminotransferase. During palmitate oxidation without aminooxyacetate or added -ketoglutarate, however, -ketoglutarate was not available for glutamate formation via glutamate dehydrogenase. We discuss the possibility that this was because (a) oxidative decarboxylation of -ketoglutarate to form succinyl-CoA was favored over glutamate formation for the competition for -ketoglutarate in the same pool, and (b) the pool of -ketoglutarate produced in the aspartate aminotransferase reaction did not serve as substrate for glutamate formation.     

Corrosion of Pembina crude oil pipeline

Summary Corrosion failure of the Pembina pipeline system of North Central Alberta, Canada, was frequent and was associated with constant bacterial load and sulphide in the crude oil and produced water. The bacterial load included a variety of anaerobic and aerobic/facultative bacteria which acted in concert to produce sulphide, giving rise to a cascade of sulphide generation.A total of 256 isolates from the crude oil were tested for ability to reduce oxidized sulphur compounds to sulphide. Five groups of bacteria, (A-E), based on this ability to reduce sulphur compounds, existed in the crude oil system. Group A reduced sulphur compounds with oxidation states +6; and lower, Group B reduced oxidation state +4 and below; Group C, oxidation states +2 and lower. Group D reduced only oxidation state 0 (elemental Sulphur), while Group E could reduce no sulphur compound to sulphide. It was found that a ceiling on the reductive capability of each bacterial group was set by the oxidation state of the sulphur compounds. The result is a synergistic relationship whereby intermediate products of reductive activities of each group form the substrate for subsequent action by other groups until sulphide is produced.     

Response of NADH oxidation and growth in K-562 cells to the antitumor sulfonylurea,N-(4-methylphenylsulfonyl)-N′-(4-chlorophenyl)urea (LY181984)

Summary Growth of K-562 cells in culture is inhibited by the antitumor sulfonylureaLY181984 (N-(4-methylphenylsulfonyl)-N-(4-chlorophenyl)urea) with an ED₅₀ of about 30 M. LY181984 was shown previously to inhibit NADH oxidation by plasma membranes from HeLa cells and other sources and to influence mitochondrial oxidative phosphorylation. With K-562 cells, NADH oxidation by plasma membranes was transiently stimulated and then inhibited by LY181984. NADH oxidation by whole cells was transiently stimulated and then inhibited by 0.1 to 100 M LY181984 as well. Both the stimulations and inhibitions of activity were time-dependent. NADH oxidation by lower phase membranes depleted of plasma membranes by aqueous two-phase partition also was inhibited by micromolar and submicromolar concentrations of LY181984. Inhibition did not correlate with mitochondrial presence but rather with membranes that appeared to be fragments of the Golgi apparatus. The oxidation of NADH by whole cells and of plasma membranes that was inhibited by LY181984 was distinguished from mitochondrial NADH oxidation by resistance to inhibition by cyanide and by proceeding under oxygen-depleted conditions or an argon atmosphere. In contrast to the active antitumor agent LY181984, the inactive but chemically-related analog, LY181985 (N-(4-methylphenyl-sulfonyl)-N-(4-phenylurea), inhibited neither growth nor NADH oxidation with K-562 cells or cell fractions.     

Lipopolysaccharide (LPS) increases thein vivo oxidation of branched-chain amino acids in the rat: A cytokine-mediated effect

Septic rats (as induced by cecal puncture and ligation) showed an increased rate ofin vivo leucine oxidation as measured from the formation of¹⁴CO₂ from an intravenously injected [1-¹⁴C]leucine tracer dose. Acute lipopolysaccharide (LPS) administration (1 mg/kg) to rats caused a similar effect on the rate ofin vivo leucine oxidation. Additionally, both tumour necrosis factor- (TNF) and interleukin-1- (IL-1), in an acute dose of 100 g/kg, also increased the rate of the oxidation of the amino acid, although only IL-1 caused a similar increase to that observed following LPS. The observed increased leucine oxidation was related to lower leucine concentrations both in LPS- and cytokine-treated rats. Important decreases were also observed in the other branched-chain amino acids (valine and isoleucine) in the LPS- and IL-1-treated animals. Isolated incubated muscles from TNF- and IL-1-treated rats did not show any changes in the rate of leucine utilization, thus suggesting that the mechanism by which the cytokines stimulate whole-body leucine oxidation is not based on an increase in the activity of the enzymatic machinery responsible for leucine oxidation. Additionally, glucocorticoids do not seem to mediate the enhancedin vivo oxidation of the amino acid since, although they are increased by both LPS and cytokines, treatment of the animals with RU486 (a glucocorticoid antagonist) was not able to suppress the effects of the cytokine onin vivo leucine oxidation.     

Cooxidation of naphthalene and other polycyclic aromatic hydrocarbons by the nitrifying bacterium,Nitrosomonas europaea

The soil nitrifying bacterium Nitrosomonas europaea has shown the ability to transform cometabolically naphthalene as well as other 2- and 3-ringed polycyclic aromatic hydrocarbons (PAHs) to more oxidized products. All of the observed enzymatic reactions were inhibited by acetylene, a selective inhibitor of ammonia monooxygenase (AMO). A strong inhibitory effect of naphthalene on ammonia oxidation by N. europaea was observed. Naphthalene was readily oxidized by N. europaea and 2-naphthol was detected as a major product (85%) of naphthalene oxidation. The maximum naphthol production rate was 1.65 nmole/mg protein-min in the presence of 240 M naphthalene and 10 mM NH₄ ⁺. Our results demonstrate that the oxidation between ammonia and naphthalene showed a partial competitive inhibition. The relative ratio of naphthalene and ammonia oxidation, depending on naphthalene concentrations, demonstrated that the naphthalene was oxidized 2200-fold slower than ammonia at lower concentration of naphthalene (15 M) whereas naphthalene was oxidized only 100-fold slower than ammonia oxidation. NH₄ ⁺- and N₂H₄-dependent O₂ uptake measurement demonstrated irreversible inhibitory effects of the naphthalene and subsequent oxidation products on AMO and HAO activity.     

Patterns of pyrite oxidation by different microorganisms

The bacterial-chemical oxidation of natural pyrites with different physical, chemical, and electrophysical characteristics by bacteria Acidithiobacillus ferrooxidans, Sulfobacillus thermotolerans, and the archaeon Ferroplasma acidiphilum were studied. The electrophysical characteristics of three natural pyrites differed in the K _thermoEMF value (pyrites 3, 4, hole conduction (p-type conductivity); pyrite 5, mixed type conductivity (n-p)) and in the logarithm of electric resistance. Chemical oxidation of pyrites 3 and 5 resulted in no changes of K _thermoEMF. When pyrite 4 was oxidized chemically, the K _thermoEMF values remained in the same range as in the initial sample, but the ratio of grains with different K _thermoEMF values in the sample was changed: the number of grains with a higher K _thermoEMF value increased. The same changes were also observed in the course of bacterio-chemical oxidation of pyrite 4. Of the three pyrites studied, an increase in the logarithm of resistance was observed only for chemical oxidation of pyrite 4 at 28°C. At higher experimental temperatures, the logarithm of resistance increased accordingly; more active bacterial-chemical oxidation resulted in a more pronounced increase in the logarithm of resistance than chemical oxidation. On bacterial-chemical oxidation of pyrites 3 and 5 by A. ferrooxidans and S. thermotolerans strains, iron was leached more actively than sulfur. Preferred bacterial-chemical oxidation of certain fractions from the pyrite samples was shown, namely of the pyrite 3 fraction with higher K _thermoEMF values by the F. acidiphilum strain and of a fraction from the pyrite 5 sample with medium K _thermoEMF values by the A. ferrooxidans and S. thermotolerans strains. The comparative assessment of bacterial-chemical pyrite oxidation by three types of microorganisms showed the direction of changes in the K _thermoEMF values to be the same in the case of bacteria Acidithiobacillus ferrooxidans and Sulfobacillus thermotolerans and different in the case of the archaeon Ferroplasma acidiphilum.     

Cytochrome c oxidation by the electron transport fraction of Azotobacter vinelandii

The spectrophotometric oxidation of horse heart ferrocytochrome c was examined by use of the particulate electron transport fraction (R(3)) of Azotobacter vinelandii strain O. Unlike cytochrome c, purified preparations of native Azotobacter cytochromes c(4) + c(5) were oxidized only slowly by the electron transport fraction. The oxidation of mammalian cytochrome c proceeded at an appreciable rate and displayed "apparent" first-order kinetics at a pH optimum of 9.0 with tris(hydroxymethyl)aminomethane-chloride buffer. The calculated V(max) value was 0.22 mumole of cytochrome c oxidized per min per mg of protein (25 C) and a K(m) value for cytochrome c of 2.3 x 10(-5)m was obtained. Ferricytochrome c was a "strict" competitive inhibitor for this oxidation. Cytochrome c oxidation by the Azotobacter electron transport system was markedly sensitive to cyanide, azide, and hydroxylamine, although carbon monoxide inhibition could not be demonstrated. It was sensitive also to high concentrations of phosphate, ethylenediaminetetraacetate, and some metal cations. "Aging" or prolonged storage of the Azotobacter R(3) fraction, at 4 C for 10 days, resulted in a threefold increase in specific activity. The cytochrome c peroxidase type of reaction did not occur with the R(3) electron transport fraction.