Role of silicon in diatom metabolism

1. In silicic acid-starved cells of the diatom Nitzschia alba, ⁶⁸Ge(OH)₄ is transported against a concentration gradient, leading to intracellular concentrations of germanic acid up to 3500 times greater than the exogenous concentrations. The accumulated substrate is osmotically active, as determined by its efflux into germanic acid-free medium.
2. Metabolic energy is required for Ge(OH)₄ transport, since uptake is completely inhibited by 1 mM DNP, 5×10^-2 M sodium azide or 1 mM iodacetamide, and is strongly inhibited by CCCP and antimycin A. Inhibition of protein synthesis with 20 μg/ml cycloheximide does not affect the initial velocity of transport, but strongly reduces the steady state intracellular concentration.
3. A double reciprocal plot of uptake velocity versus substrate concentration yields a biphasic curve. The kinetic data are consistent with the interpretation that N. alba has two transport systems for germanic acid; a high affinity-low capacity (K _s=0.36 μM; V _max 1.2 μmoles/10⁸ cells/min) system and a low affinity-high capacity (K _s=5 μM; V _max 6.2 μmoles/10⁸ cells/min) system.
4. The implications of these findings for silicic acid transport and metabolism in N. alba are discussed.

     

Uptake of guanine by synchronizedChlorella fusca

1. The transport of guanine in autospores of light-dark synchronizedChlorella fusca was studied using radioactive guanine in the concentration range of 4 nM to 50 μM.
2. The transport system was constitutive, it had high specificity for the permeant, and theQ ₁₀ value was in the range of 1.5 to 2.2. At concentrations lower than 0.2 μM the half saturating constant, S_0.5 was 1 μM both for cells kept in dark and cells kept in light. At higher concentrations the S_0.5 of darkened cells was about 0.23 μM, while that of illuminated cells was unchanged. Only above 0.2 μM guanine did illumination of the cells or addition of glucose increase the transport rate.
3. Guanine which had accumulated did not leak out at temperatures below 45°C or by treatment with 10 μM dinitrophenol, which completely inhibited transport. Furthermore, the accumulated guanine did not exchange with exogenous guanine.
4. The guanine accumulated, more than 10⁵-fold over the external concentration, showing that the transport, was active.
5. The initial transport rate per cell revealed annual fluctuations.

     

δ-Aminolevulinic acid transport in Saccharomyces cerevisiae

• 1.1. This work represents the first approach to characterize the transport system of haem pathway precursors, such as δ-aminolevulinic acid (ALA), in two strains of Saccharomyces cerevisiae, a wild type, D27, and a HEM R⁺ mutant.
• 2.2. ALA transport occurs unidirectionally by a sole active system with an apparent K_M of 0.10 mM, at the optimum pH of 5.0. ALA uptake is influenced by both the carbon and nitrogen source; this suggests a rather complex regulation mechanism.
• 3.3. This transport is not mediated by the general amino acid permease (GAP).
• 4.4. ALA uptake is strongly inhibited by compounds harboring a methyl-amine terminus suggesting that this group is essential for ALA transport; however, the electric environment of the carboxylic group may be also important for the interaction between ALA and its transporter active site.
• 5.5. We have found differences in ALA transport which would indicate a different regulation mechanism for this system in both strain cells.

     

EPR characterization of oxide supported transition metal ions: Relevance to catalysis

EPR spectroscopy is a powerful tool to identify at a molecular level, the different steps of catalyst preparation, and of catalytic reactions:

1. Deposition of paramagnetic transition metal ions onto a support is monitored, and the coordination sphere of the metallic center is characterized by EPR.
2. The catalyst is also characterized after activation (thermal oxidation or reduction):

- the distribution among the different sites in zeolites can be determined;

- the dispersion of the active phase may be appreciated;

- the unsaturation degree of the active site may be evaluated using probe molecules such as water or¹³C enriched carbon monoxide.

1. The catalytic mechanisms can be investigated by studying the elementary steps of the catalytic reaction, as illustrated for methanol oxidation over Mo/SiO₂ catalysts whose EPR results have extended the reaction mechanism proposed on the basis of kinetic data. In addition, reaction intermediates may be isolated inquasi-in situ conditions as in the case of olefin oligomerization catalyzed by Ni/SiO₂ systems.

     

Modulation of active potassium transport by aldosterone

• 1.1. The role of aldosterone on active potassium transport across lizard colon under voltage-clamped conditions has been investigated.
• 2.2. Control colons exhibited no net potassium flux (J^k_net) despite of the existence of active opposite unidi ectional fluxes.
• 3.3. An important net secretory potassium flux was found in short-circuited aldosterone-stimulated colons.
• 4.4. Mucosal amiloride did not change (J^k_net) either in control or aldosterone-stimulated colons.
• 5.5. Luminal barium alters K ⁺ transport in a manner consistent with the presence of barium-sensitive conductances at the apical membrane of both control and aldosterone-treated colons.
• 6.6. The effects of ouabain and barium on control and aldosterone-induced potassium flows were consistent with a model involving basolateral uptake by an Na ⁺-K ⁺-ATPase and conductive exit across the apical membrane.
• 7.7. The stimulatory effect of aldosterone on potassium secretion is associated with parallel increases of both basolateral K ⁺ entry and the apical conductive pathway.

     

Growth of a floating aquatic weed,Salvinia under standard conditions

1. Growth of the floating aquatic weed, Salvinia, in sterile culture was exponential for at least 2 weeks under standardized conditions.
2. Increase in light intensity or in CO₂ resulted in increases in growth rate, but did not extend the exponential period of growth.
3. This aquatic plant, like many others, discriminates against calcium relative to strontium.
4. In culture Salvinia exhibited luxury consumption of N and P.
5. Because of high C/N ratios, Salvinia may not be a favorable source of animal food, but might be useful in nutrient removal schemes.
6. In sterile culture, S. molesta produced fewer leaves than S. minima, but maintained a significant increase in leaf area and dry weight. This may be correlated with the ability of the first species to rapidly spread over tropical waterways.

     

On the possible role of respiratory activity of Acholeplasma laidlawii cells in sugar transport

1. Out of 20 exogeneous substrates only ethanol and, to a much lesser extent, lactate and pyruvate were shown to be capable of stimulating the respiration of Acholeplasma laidlawii cells. However, none of these substrates changed the initial rate of active transport of 3-O-methyl-d-glucose (3-O-MG).
2. From inhibitory analyses and spectroscopic data, it is apparent that the respiratory chain of A. laidlawii has no cytochromes and is probably not responsible for oxidative phosphorylation.
3. Valinomycin and nigericin stimulated cell respiration only in the presence of K⁺-ions, while monensin stimulated it in the presence of Na⁺-ions.
4. 3-O-MG transport was shown to be sensitive to uncouplers, ATPase inhibitors and arsenate are resistant to a majority of respiratory inhibitors tested. This suggested that there was no relationship between respiration and carbohydrate transport in the A. laidlawii cells. Further evidence was provided by the absence of respiratory stimulation during the transport of non-metabolizing carbohydrates.

     

Rationalization of properties of nitrate reductases in Rhodopseudomonas capsulata

1. The properties of nitrate reductase activities have been compared in several strains of Rhodopseudomonas capsulata grown phototrophically in the presence of nitrate as sole nitrogen source.
2. Strains AD2 and BK5 resemble the spontaneous mutant N22DNAR⁺ (described by McEwan et al. 1982 FEBS Lett. 150, 277\2-280) in that reduction of nitrate was inhibited by either illumination or oxygen but not by NH ₄ ⁺ , and that electron flow to nitrate under dark anaerobic conditions generated a cytoplasmic membrane potential (as judged by an electrochromic shift in the absorbance spectrum of endogenous carotenoid pigments). In contrast disappearance of nitrate from suspensions of strains N22 and St. Louis was dependent upon illumination and was inhibited by NH ₄ ⁺ . Membrane potentials were not generated by addition of nitrate in the dark to N22, St. Louis or strain Kbl.
3. Nitrate reductase was shown to be located in the periplasmic space of both strain AD2 and mutant N22DNAR⁺. The nitrate reductase activity in cells of AD2 and N22DNAR⁺ was relatively insensitive to azide, with 0.5mM azide required for 50% inhibition. The nitrate reductase of strain BK5 was more strongly associated with the cytoplasmic membrane and no conclusion could be reached about whether it was located on the periplasmic or cytoplasmic surface. In BK5 cells nitrate reductase activity was sensitive to low concentrations of azide (50% inhibition with 2 \gmM azide). It is proposed that functionally the nitrate reductase activity in strains AD2, BK5 and N22DNAR⁺ has identical roles. These roles are suggested to include:
4. The first step in the assimilation of nitrate.
5. Provision of an alternative electron acceptor to oxygen for generating a membrane potential.
6. A mechanism for disposing of excess reducing equivalents in the maintenance of balanced growth. This type of nitrate reductase, especially in AD2 and N22DNAR⁺, appears to resemble that described in a denitrifying strain of Rps. sphaeroides, but to differ markedly from its membrane-bound counterpart in other bacteria including the denitrifying Paracoccus denitrificans and Escherichia coli.
7. In other strains of Rps. capsulata including St. Louis, N22 and Kbl, only an assimilatory nitrate reductase, whose activity in intact cells is relatively sensitive to azide, is present in anaerobic, phototrophic cultures grown with nitrate as nitrogen source. As this reductase cannot be detected after breakage of cells, no conclusion can be made as to its location in the cell.

     

Reverse electron flow in chloroplasts

Energy dependent reverse electron flow reactions in isolated thylakoids provide a unique tool to study, in the dark, the coupling between the ATP synthase, proton transport and the electron transfer system. Appropriate experimental conditions have been established to follow experimentally the following reactions:

1. ATP driven proton uptake into the inner-thylakoid space, which requires preactivation of the ATP synthase.
2. ATP driven reverse electron transport, which involves proton transport as an intermediate, and results in the reduction of Q_A by an externally added electron donor.
3. ATP driven luminescence, which requires the presence of an oxidized partner on the water side of photosystem II, and involves electron transport from Q_B to Q_A.
4. ΔpH driven reverse electron flow, which does not require the participation of the ATP synthase, and uses reduced intermediates between the two photosystems as electron donors for the reduction of Q_A.
5. ΔpH driven luminescence which again uses reduced intermdiates between the two photosystems as electron donors for Q_A reduction, and requires the presence of an oxidized partner on the water side of photosystem II.

Several of these reactions have been shown to occur in intact chloroplasts and may provide an important regulatory mechanism in vivo.     

The salt relations of marine and halophilic species of the unicellular green alga,Dunaliella

1. Comparisons were made of the effects of salt on the exponential growth rates of two unicellular algae,Dunaliella tertiolecta (marine) andDunaliella viridis (halophilic).
2. The algae contained glycerol in amounts which varied directly with the salt concentration of the growth media. The highest measured glycerol content ofD. tertiolecta was approximately equivalent to 1.4 molal and occurred in algae grown in 1.36 M sodium chloride. The highest glycerol content measured inD. viridis was approximately equivalent to 4.4 molal and occurred in algae grown in 4.25 M sodium chloride. Lower concentrations of free glucose, which varied inversely with extracellular salt concentration, were also detected.
3. It is inferred that Na⁺ is effectively excluded from the two algae. There was some evidence of a moderate uptake of K⁺.
4. Comparisons were made of erude preparations of the glucose-6-phosphate dehydrogenase and an NADP-specific glycerol dehydrogenase from each species and of the effects of salt and glycerol on the activities of these enzymes. It is concluded that the different salt tolerances of the two algae cannot be explained by generalized differences between their enzyme proteins.
5. Although intracellular glycerol must necessarily contribute to the osmotic status of the algae, its primary function in influencing their salt relations is considered to be that of a compatible solute, whereby glycerol maintains enzyme activity under conditions of high extracellular salt concentration and hence low (thermodynamic) water activity.

     

Effects of in vivo ethanol administration on Ca2+/Mg2+ ATPase and ATP-dependent Ca2+ uptake activity in synaptosomal membranes

1. Acute administration of ethanol (4 g/kg, i.p.) to mice inhibits the sequestration of calcium into endoplasmic reticulum-like organelles in synaptosomal membranes.
2. Ethanol administration inhibits both Ca²⁺-stimulated adenosine triphosphate hydrolysis and ATP-dependent calcium uptake in the vesicles at time of loss of righting reflex.
3. At recovery of righting reflex, the Ca²⁺-ATPase activity returns to normal levels, while the ATP-dependent uptake remains inhibited.
4. The effect of ethanol is specific for the sequestration (active transport) of calcium since calcium binding to synaptic membranes is not altered.
5. Alteration in mechanisms responsible for synaptosomal buffering of cytosolic Ca²⁺ levels by in vivo ethanol may contribute to altered transmitter release rates following ethanol adminstration.

     

Charakterisierung eines phagenähnlichen Partikels aus Zellen von Nitrobacter

1. Polyhedral particles were isolated from cells of Nitrobacter winogradskyi and of Nitrobacter strains K₁, K₄ and α₁. Their physical and biological properties are characterized.
2. The investigated strains contain polyhedral particles, 1000–1200 Å in size. With increasing age of the culture more particles are found in cells of Nitrobacter. Simultaneously the number of colony producing nitritoxidants decreases.
3. In strain α₁ the loss of the capability to form colonies is connected with partial lysis of the cell and release of particles.
4. A homogeneous fraction of particles was obtained by zone density gradient centrifugation in Tris-Mg-SH-buffer.
5. The polyhedral particles have a sedimentation coefficient of s _w,20 ⁰ =825S and a CsCl-buoyant density of ?²⁵ g/cm³.
6. Based on the determined properties the particles are classified as phage-like Nitrobacter particles Nb₁.

     

Cyclic AMP stimulation and prostaglandin inhibition of Na transport in freshwater mussels

• 1.1. Pondwater acclimated Carunculina texasensis and Ligumia subrostrata experienced a 230% increase in Na influx when injected with dibutyryl cyclic AMP (0.4mM/l blood).
• 2.2. Theophylline, a phosphodiesterase inhibitor, or indomethacin, an inhibitor of prostaglandin (PG) synthetase, caused a dose dependent stimulation of Na transport.
• 3.3. Prostaglandin E₂ injected into mussels caused an inhibition of Na influx. Arachidonic acid, the precursor of PGE₂, inhibited Na influx or stimulated Na efflux depending on the animal's acclimation conditions.
• 4.4. Chloride transport was unaffected by the drugs used in this study.

     

The effect of inhibitors on the electron flow triggering photo-phobic reactions in cyanophyceae

1. Since photo-phobic reactions in the blue green alga Phormidium uncinatum seem to be triggered by changes of electron flow rates into or out of an electron pool situated in the electron transport chain between photosystem II and I, the effect of inhibitors affecting the electron transport chain has been studied.
2. Dose response curves of the phobic reaction have been measured by varying the trap energy in double beam light trap experiments with constant pairs of monochromatic light. From these dose response curves the effects of the inhibitors on both types of phobic reactions, i.e. exit reactions and entrance reactions, have been calculated.
3. Dibromothymoquinone (DBMIB) inhibits the electron transport between the electron pool and photosystem I by preventing the reoxidation of plastoquinone. The phobic entrance reaction, which results in an emptying of the light trap, is triggered by changes in the electron flow out of the pool; thus it is more effected by DBMIB than the exit reaction, which is mediated by the electron transport into the pool.
4. The phobic exit reaction, which results in accumulations in the light trap, is triggered by changes in the electron flow into the electron pool via photosystem II. 3-[3,4-dichlorophenyl]-1,1-dimethylurea (DCMU) inhibits the electron transport near photosystem II; thus it affects the exit reaction more than the entrance reaction.

     

Phosphorylative fumarate reduction in Vibrio succinogenes: Stoichiometry of ATP synthesis

1. Cells of Vibrio succinogenes, treated with EDTA at pH 8, catalyze the phosphorylation of their endogenous ADP and AMP as a function of the electron transport from formate to fumarate. The P/fumarate ratio obtained from the initial velocity of the phosphorylation on initiation of the electron transport and from the activity of fumarate reduction in the steady state was 0.90. The phosphorylation was prevented by 10μmol/g protein carbonylcyanide-3-chlorophenylhydrazone.
2. The esterification of external phosphate in the presence of ADP, hexokinase and glucose is catalysed by a membrane preparation of V. succinogenes in the steady state of fumarate reduction by H₂. The phosphorylation was fully abolished by either 5μmol/g protein carbonylcyanide-4-trifluoromethoxyphenylhydrazone or 30μmol/g protein carbonylcyanide-3-chlorphenylhydrazone. Phosphorylation was blocked also by dicyclohexylcarbodiimide, an inhibitor of the Mg²⁺-dependent membrane bound ATP synthase, and by low concentrations of the inhibitors of electron transport 2-(n-nonyl)-4-hydroxyquinoline-N-oxide or 4-chloromercuriphenylsulfonate.
3. The P/fumarate ratios, measured with the membrane preparation, were found to increase with progressive inhibition of the electron transport from hydrogen to fumarate by means of 4-chloromercuriphenylsulfonate. The extrapolated ratio at vanishing electron transport activity was 0.47.
4. About 50% of the membrane preparation was found to consist of inverted vesicles with the hydrogenase and formate dehydrogenase oriented to the inside. The residual part is considered as being incapable of performing energy transduction. The extrapolated P/fumarate ratio valid for the inverted vesicles was 0.94.

     

Somatic embryogenesis and plant regeneration in garden leek

High frequency plant regeneration via somatic embryogenesis has been induced from in vitro shoot-base cultures of seedlings of garden leek (Allium porrum L.). Four main steps are involved in the procedure using BDS medium:

- shoot multiplication with 17.6 mM benzyladenine;

- induction of nodular callus from the in vitro shoot base with 9 mM 2,4-dichlorophenoxyacetic acid;

- initiation of embryogenic callus from nodular callus with 9 mM 2,4-dichlorophenoxyacetic acid +7.6 mM abscisic acid;

- plant regeneration from embryogenic callus with 9.8 mM N⁶-(2-isopentenyl)adenine.

The presence of 2,4-dichlorophenoxyacetic acid in the medium and light conditions were shown to be essential for nodular callus induction and somatic embryogenesis. Abscisic acid was not a prerequiste for somatic embryogenesis, but it significantly increased the frequency.     

Proton translation in submitochondrial vesicles

An investigation of proton translocation in submitochondrial vesicles from rat liver has been made under simple experimental conditions. Choline chloride was used both as the oxidizable substrate and the ionic medium for the measurement of activity during oxygen pulse experiments:

1. The passive permeability measured from the decay of proton efflux after an oxygen pulse could be described by a first-order equation. An H⁺/O ratio of 2·5 was obtained for choline oxidation in the presence of oligomycin and/or MgCl₂. Oligomycin decreased the passive proton permeability and respiration, concomitant with an increase in proton uptake. Respiratory control was directly related to the passive proton permeability and inversely related to the magnitude of the proton gradient. The decreased respiration and passive permeability reflecting respiratory control is most evident in the pH rang 5·8–7·5.
2. Preparation of submitochondrial vesicles in the presence of EDTA resulted in proton production during an oxygen pulse given at alkaline pH. Cytochromec enhanced proton uptake by approximately 1 H⁺/cytochromec, but only in the presence of Triton X-100. These results are indicative of the asymmetric behavior of the coupling membrane and provide direct evidence of the participation of electron transport components in proton translocation.

     

Neutral amino acid transport in an androgen-dependent epithelium

• 1.1. Transport of neutral amino acids by the isolated seminal vesicle epithelium of normal and gonadectomized guinea pigs has been investigated by measurement of the uptake of 2-amino[1-¹⁴C]-isobutyric acid and 2-methylamino[1-¹⁴C]isobutyric acid.
• 2.2. The V_max for Na-dependent and -independent transport of both amino acids was reduced by gonadectomy but the general transport characteristics appeared to be unchanged by this treatment.
• 3.3. The most likely explanation of the decreased transport is the loss of transporter molecules associated with the tissue regression that follows rapidly on gonadectomy.

     

Reabsorption and accumulation of α-amino-iso-butyric acid in the nephridia of Sabella pavonina Sa vigny (Annelida polychaeta)

• 1.l. High amino acid concentrations were found in the anterior coelomic fluid of a Polychaeta (Sabella pavonina Savigny).
• 2.2. The concentrations being much higher in the fluid which penetrates the nephrostomia into the nephridia lumen than in the final urine indicates that the nephridia reabsorbs large amounts of amino acids.
• 3.3. Nephridial perfusion experiments showed that an amino acid analogue (α-amino-iso-butyric acid, AIB) is transported by the nephidia.
• 4.4. The transport took place across the nephridial wall owing to the presence of a carrier-mediated transport system and a diffusion system.
• 5.5. For the carrier-mediated transport, the V_max was 0.234 ± 0.025 nmol·min⁻ and the K_m 3.715 ± 0.315mmol·l⁻.
• 6.6. AIB accumulated in the nephridial cells up to a maximum rate of 01.17 nmol·min⁻.
• 7.7. Intracellular accumulation stopped increasing when the V_max for reabsorption was reached.
• 8.8. These results indicate that the carrier-mediated transport of AIB is located at the apical membrane of the nephridial cell, and that AIB transport by simple diffusion takes place through the paracellular pathway.

     

The effects of heavy metals and metabolic inhibitors on calcium uptake by gills and scales of Fundulus heteroclitus in vitro

• 1.1. Cadmium (Cd) and zinc (Zn) were inhibitory to calcium uptake by isolated gills of Fundulus heteroclitus in vitro. The metals appeared to act by displacing Ca²⁺ ions from protein carriers involved in facilitated diffusion.
• 2.2. In saltwater fish, transport of calcium across the serosal membrane of gill chloride cells is partly energy dependent and is likely mediated by Ca²⁺-ATPase. However, much of the calcium transport through the gill epithelium appears to occur by passive processes.
• 3.3. Cd (10⁻⁵M—10⁻³M) and Zn (10⁻⁷M—10⁻³ M) inhibited calcium uptake by isolated scale patches incubated in a physiological saline.
• 4.4. Cyanide, oubain, and quercetin treatment of scale patches produced results similar to those of the Cd and Zn treatments suggesting that metal-induced inhibition of ATPases may be responsible for reduced calcium transport by scale osteoblasts.