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.

     

Acid-base titration across the plasma membrane ofMicrococcus denitrificans: Factors affecting the effective proton conductance and the respiratory rate

The object of this work was to measure the effective proton conductance of the plasma membrane ofMicrococcus denitrificans under various conditions and to investigate possible connections between respiration and proton translocation.

1. Pulsed acid-base titrations of suspensions ofM. denitrificans in a medium containing the permeant thiocyanate ion, or when K⁺ ion permeability was induced by valinomycin in a KCl medium, showed that the normal effective proton conductance of the membrane system was less than 1 μmho/cm².
2. A pH-overshoot artefact was suppressed by adding carbonic anhydrase.
3. The effective proton conductance was increased by the uncoupler FCCP in the same concentration range as was required to stimulate respiration. Concentrations of FCCP above 1·5 μM inhibited respiration after an initial stimulation.
4. The effective proton conductance in presence of 2 μM FCCP was at least 17 μmho/cm².
5. The quantitative relationships between the respiratory rate, the stoichiometry of respiration-driven proton translocation, and the effective proton conductance of the membrane of the cells are compatible with the suggestion that stimulation of respiration by FCCP is due to a release of back-pressure exerted by a protonmotive potential on the respiratory chain system in the membrane. Only one amongst other possible explanations of the stimulation of respiration by FCCP is, however, excluded.

     

The role of mitochondria in modifying the cellular inoc environment: Studies of the kinetic accumulation of calcium by rat liver mitochondria

1. The affinity of ATP-supported Ca²⁺ accumulation for both Ca²⁺ and ATP was determined from initial rate studies employing isolated rat liver mitochondria. TheK _m values for “free” Ca²⁺ and ATP were calculated to be of the order of 2 μM and 100 μM, respectively. TheK _m for ATP decreased as the Ca²⁺ concentration was increased.
2. The curve relating initial rates of Ca²⁺ accumulation to Ca²⁺ concentration was singmoidal in shape; values obtained for the Hill coefficient were in the range 1.5–1.9.
3. Concomitant with the ATP-stimulated accumulation of Ca²⁺, ATP translocation was itselt increased in the presence of Ca²⁺. This stimulation took place independently of Ca²⁺ accumulation.
4. Decreasing the pH of the incubation medium decreased the rate of Ca²⁺ accumulation. This inhibition was competitive in that the affinity of mitochondrial for Ca²⁺ could be altered. The maximal rate of accumulation did not change with change in pH.
5. The permeant anions inorganic phosphate and acetate stimulated the accumulation of Ca²⁺ in a non-competitive manner. Both theV _max and K_m varied when either of the anions were present.
6. The data are discussed in relation to the role that mitochondria play in controlling the cellular ionic environment.

     

Oxaloacetate decarboxylase from Acetobacter aceti

An oxaloacetate (OAA) decarboxylase (EC 4.1.1.3) has been purified 72-fold from Acetobacter aceti cells grown on ethanol, and its molecular weight was estimated to be about 80,000 by gel filtration. Several properties distinguished this enzyme from the OAA decarboxylase from A. xylinum:

1. It was not a constitutive enzyme; the activity was 6- to 20-fold higher in cells grown on a C₂ substrate (acetate or ethanol) than in cells grown on a C₃ compound (pyruvate or propionate).
2. The optimum pH was 7.5; a value of 5.6 was reported for the enzyme from A. xylinum.
3. The enzyme did not need a divalent cation and was not inhibited by EDTA.
4. The K _mvalue for OAA was found to be 0.22 mM. It was not affected by the addition of nicotinamide adenine dinucleotide.
5. The enzyme activity was neither inhibited by acetate nor by L-malate.

In addition, the OAA decarboxylase from A. aceti was insensitive to monovalent cations, avidin or acetyl coenzyme A.     

Adenine nucleotide pool variations in intact Nitrobacter winogradskyi cells

1. The ATP pool in Nitrobacter winogradskyi cells was determined by means of the luciferin-luciferase enzyme system and the ADP and AMP pools were measured after enzymatic conversion into ATP.
2. In the fist 10 min after addition of nitrite to endogenously respiring cells, which had stood for 5–16 days after completion of the nitrite oxidation, the ATP pool dropped about 60%.
3. During the log phase the ATP pool was approx. 20–40 pmoles/5 μg cell-N. During growth it increased exponentially by 3–4 times the amount until the nitrite had been used up. Subsequently the ATP pool decreased at first rapidly and then more slowly without sinking to 0 in the first 2 months after nitrification.
4. Nitrite oxidizing cells had an energy charge of 0.37 during the log-phase. After approx. 90% of the substrate had been used up the energy charge had reached 0.57.
5. If the CO₂ assimilation was inhibited in growing cultures by increased oxygen partial pressure, nitrite oxidation continued but the ATP pool increased.
6. The ATP pool and the activity of the endogenous respiration decreased by more than 50% during the first hours after the substrate had been used up.

     

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.

     

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.

     

Charkterisierung eines phagenähnlichen Partikels aus Zellen von Nitrobacter

1. Phage-like particles Nb₁ isolated from cells of Nitrobacter agilis were characterized after freeze etching and after treatment by fixation agents.
2. Ethanol-acetic acid fixed particles can be digested by the proteolytic enzyme papain.
3. Ethanol-acetic acid fixed particles show a loss in mass and volume after treatment with DNase. Under the same conditions RNase has no influence.
4. The chemical composition of the phage-like particle Nb₁ is discussed.

     

Inhibition of carotenoid synthesis in Myxococcus fulvus (Myxobacterales)

1. The inhibitory effects of CPTA, nicotine, DPA, and San 6706 on carotenogenesis in Myxococcus fulvus were investigated.
2. The effects of CPTA, D-nicotine, and L-nicotine were very similar. The action of the drugs wasadditive. The cyclization was inhibited at low doses, the introduction of the hydroxyl group at C-1′ at higher doses. Lycopene accumulated at high drug concentration. The mode of action of the inhibitors is discussed.
3. In a carotenoid mutant of M. fulvus a stimulation of the “7,8-dehydrogenase” by CPTA was observed.
4. The specific carotenoid content of bacteria was increased by DPA due to an enhanced formation of phytoene. At low doses of DPA small amounts of an intermediate carotenoid glucoside ester, a 7,8-dihydro derivative, were detected.
5. DPA was taken up by the plasma membrane. Quantitative removal of DPA by washing was not possible.
6. San 6706 specifically and reversibly blocked the desaturation of phytoene.

     

In vitro effects of sodium selenite on nuclear 3,5,3’-triiodothyronine (T3) receptor gene expression in rat pituitary GH4C1 cells

The present study was undertaken in order to investigate the effects of sodium selenite on:

1. The growth of rat pituitary GH₄C₁ cells;
2. The nuclear T₃ receptor gene expression;
3. The cytoplasmic protein phosphorylation; and
4. The prolactin secretion in rat pituitary GH₄C₁ cell line.

Sodium selenite (up to 2.5 μM) has no inhibitory effect on GH₄C₁ cell proliferation as well as the prolactin secretion. On the other hand, 0.5 μM sodium selenite significantly decreases the rate of mRNA synthesis and/or degradation of both, the α1 form of the T₃ receptor (TRα1) and the α2 isoform of the T₃ receptor. At 1 μM of sodium selenine, significant changes in the electrophoretic profile of low molecular mass cytoplasmic proteins were found, moreover, sodium selenite (1 μM) also considerably affects phosphorylation of a higher molecular mass proteins. The results based on the in vitro experiments suggest that sodium selenite may affect specific processes at the pretranslational level as well as it may also take part in processes of posttranslational modification of protein(s), the cell vitality and the cell growth remaining unchanged.     

Intracellular proteolytic activity in developing myxospores of Myxococcus xanthus

1. Cell-free extracts from vegetative cells and developing myxospores of Myxococcus xanthus were found to contain similar amounts of proteolytic activity, approximately 80% of which was due to one or more neutral metal proteases.
2. Sixty per cent of the proteolytic activity was particulate.
3. The specific activity of the proteases was high throughout all stages of myxospore formation and displayed small increases in activity at two stages of development: (1) during cell shortening and (2) immediately following the conversion to spheres. The first peak in activity was apparent in assays conducted at pH 8 or 10 whereas the second peak was obvious only at pH 6.
4. A mutant which develops into myxospores only after a lag of approximately 7–8 h possessed levels of proteases similar to the wild type and displayed a peak in proteolytic activity after a delay of 7–8 h.
5. Low levels of serine protease activity were occasionally detected in both vegetative cells and myxospores; no sulfhydryl proteases were detectable in either cell type.
6. Extracellular proteases accumulated in the medium throughout myxospore development but differed from the intracellular proteases in pH optima and sensitivity to inhibitors.

     

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.     

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.

     

Growth of Nitrobacter in the presence of organic matter

1. Culture filtrates of heterotrophic bacteria were tested for their stimulatory effect on nitrification of three strains of Nitrobacter.
2. Yeast extract-peptone solution, in which Pseudomonas fluorescens had grown, after removal of the cells was added to autotrophically growing cultures of Nitrobacter agilis; it caused a stimulated nitrite oxidation and growth of Nitrobacter agilis.
3. The degree of stimulation depended on: a) the proportion of the culture filtrate to the autotrophic medium; b) the composition of the complex medium in which Pseudomonas fluorescens had been grown; c) the time the heterotrophic bacterium had been grown in the complex medium.
4. The stimulatory effect was highest with Nitrobacter agilis, less with Nitrobacter winogradskyi and negligible with Nitrobacter K ₄.
5. It was possible to adapt nitrifying cells of Nitrobacter agilis to higher concentrations of yeast extract and peptone. After the nitrite had been completely oxidized the cell-N still increased up to 30% before growth stopped.

     

Coordination of energy and carbon metabolism in lysine producing Brevibacterium strains

The present paper deals with the coordination of energy metabolism, glucose consumption rate, glycolytic and TCA cycle enzyme activities in the lysine-producing bacterium Brevibacterium flavum. It is shown, that inhibition of the elctron transport chain causes changes of the following sequence:

at first, TCA cycle enzymes are activated;

secondly, TCA cycle enzyme activity decreases, and glycolytic enzyme activities as well as glucose transport rate increase; there is a slight increase in Q_o2 and a considerable one of O₂ consumption in cyanide-resistant respiration pathway;

thirdly, TCA cycle enzyme activities and glucose transport rate decrease.

It is supposed, that coordination of carbon and energy metabolism in B. flavum depends on intracellular ATP concentration or energy charge value.     

Chlorophenol and chlorobenzoic acid co-metabolism by different genera of soil bacteria

1. FourPseudomonas strains, anAchromobacter sp., threeNocardia strains,Mycobacterium coeliacum and aBacillus sp., grown on either phenol or benzoic acid as sole carbon source, were tested for co-oxidation of monochlorophenols and monochlorobenzoates respectively.
2. ThePseudomonas spp.,Nocardia spp.,M. coeliacum and theBacillus sp. showed O₂-uptakes in the presence ofo-, m- orp-chlorophenol and oxidised them to either 3- or 4-chlorocatechol.
3. Benzoate-grown cells of the threeNocardia spp. and three of the pseudomonads absorbed O₂ only in the presence ofm-chlorobenzoate, 4-chlorocatechol being identified as the oxidation product in two cases. One pseudomonad gave some O₂-uptake also in presence ofp-chlorobenzoate.
4. Benzoate-grownBacillus cells gave O₂-uptakes with eitherm- orp-chlorobenzoate and the former was oxidised to 5-chloro-2,3-dihydroxybenzoic acid via 5-chlorosalicylic acid, thus representing a novel metabolic pathway.

     

Fumarate reduction inProteus mirabilis

1. Proteus mirabilis formed fumarate reductase under anaerobic growth conditions. The formation of this reductase was repressed under conditions of growth during which electron transport to oxygen or to nitrate is possible. In two of three tested chlorateresistant mutant strains of the wild type, fumarate reductase appeared to be affected.
2. Cytoplasmic membrane suspensions isolated from anaerobically grownP. mirabilis oxidized formate and NADH with oxygen and with fumarate, too.
3. Spectral investigation of the cytoplasmic membrane preparation revealed the presence of (probably at least two types of) cytochromeb, cytochromea ₁ and cytochromed. Cytochromeb was reduced by NADH as well as by formate to approximately 80%.
4. 2-n-Heptyl-4-hydroxyquinoline-N-oxide and antimycin A inhibited oxidation of both formate and NADH by oxygen and fumarate. Both inhibitors increased the level of the formate/oxygen steady state and the formate/fumarate steady state.
5. The site of inhibition of the respiratory activity by both HQNO and antimycin A was located at the oxidation side of cytochromeb.
6. The effect of ultraviolet-irradiation of cytoplasmic membrane suspensions on oxidation/reduction phenomena suggested that the role of menaquinone is more exclusive in the formate/fumarate pathway than in the electron transport route to oxygen.
7. Finally, the conclusion has been drawn that the preferential route for electron transport from formate and from NADH to fumarate (and to oxygen) includes cytochromeb as a directly involved carrier. A hypothetical scheme for the electron transport in anaerobically grownP. mirabilis is presented.

     

A study of the relationship between mechanical characteristics and the coastal vegetation among several broad-leaf trees in Miura Peninsula in Japan

The mechanical properties of broad-leaf tree species in a maritime-wind exposed habitat in central Japan were examined. The broad-leaf trees studied were Celtis sinensis var. japonica, Ilex integra, Eurya japonica, Pittosporum tobira, Euonymus japonicus and Cinnamomum japonicum. The results obtained can be summarized briefly as follows:

1. At places with weaker wind, the number of species increased and the height of the canopy increased.
2. The fracture strength σ_m showed no dependence on tree part or branch thickness, but was constant.
3. The order of strength was Celtis sinensis var. japonica > Ilex integra > Eurya japonica > Pittosporum tobira > Euonymus japonicus > Cinnamomum japonicum, and these six species could best adapt to the wind pressure in the study area.
4. Within species, fracture strength varied directly with wind strength.
5. The strain ε_m decreased as the trunk became thicker.
6. Within species, strain energy U_m varied directly with wind strength.

     

Chemolithotrophic growth and regulation of hydrogenase formation in the coryneform hydrogen bacterium strain 11/x

1. The present paper deals with the chemolithotrophic growth of a Gram-positive hydrogen bacterium strain 11/x which shows the characteristic features of some coryneform bacteria.
2. Like other hydrogen bacteria, the strain 11/x is a facultative chemolithotroph and grows on many organic substrates faster than in a mineral medium under an atmosphere of knallgas+CO₂. Fully induced, autotrophically grown cells, subcultured mixotrophically on fructose show additive growth.
3. Cell-free extracts of autotrophically grown cells are able to reduce methylene blue, dichlorophenolindophenol, phenazine methosulphate, menadione, and FMN with hydrogen. Conditions for direct NAD(P) reduction could not be found.
4. Hydrogenase is formed under autotrophic as well as mixotrophic conditions. In the latter case the rate of hydrogenase formation is diminished depending on the organic substrate. Heterotrophically grown cells do not have any detectable hydrogenase activity. For the induction of hydrogenase in those cells a nitrogen source is a prerequisite.
5. The formation of ribulose-1,5-diphosphate carboxylase and phosphoribulokinase seems to be regulated in a way similar to that of hydrogenase: the enzymes could only be detected in autotrophically and mixotrophically grown cells but not in those grown heterotrophically.

     

The anaerobic metabolism of malate of Saccharomyces bailii and the partial purification and characterization of malic enzyme

1. The main pathway of the anaerobic metabolism of l-malate in Saccharomyces bailii is catalyzed by a l-malic enzyme.
2. The enzyme was purified more than 300-fold. During the purification procedure fumarase and pyruvate decarboxylase were removed completely, and malate dehydrogenase and oxalacetate decarboxylase were removed to a very large extent.
3. Manganese ions are not required for the reaction of malic enzyme of Saccharomyces bailii, but the activity of the enzyme is increased by manganese.
4. The reaction of l-malic enzyme proceeds with the coenzymes NAD and (to a lesser extent) NADP.
5. The K _m-values of the malic enzyme of Saccharomyces bailii were 10 mM for l-malate and 0.1 mM for NAD.
6. A model based on the activity and substrate affinity of malic enzyme, the intracellular concentration of malate and phosphate, and its action on fumarase, is proposed to explain the complete anaerobic degradation of malate in Saccharomyces bailii as compared with the partial decomposition of malate in Saccharomyces cerevisiae.