Showing papers by "University of Konstanz published in 1976"

Maltose Transport in Escherichia coli K12

Abstract: The kinetic parameters for the maltose transport system in Escherichia coli K12 were determined with maltose and maltotriose as substrates. The system exhibits an apparent Km of 1 muM for maltose and 2 muM for maltotriose. The V of entry was determined as 2.0 and 1.1 nmol substrate/min per 10(8) cells. Mutations in lamB, the structural gene for the receptor protein of phage lambda, increased the Km for maltose transport by a factor of 100-500 without influencing the maximal rate of transport. Maltotriose is no longer transported in these lamB mutants. The maltose-binding protein, an essential component of the maltose transport system, was found to exhibit substrate-dependent fluorescence quenching. This phenomenon was used to determine dissociation constants and to estimate the rate of ligand dissociation. A Kd of 1 muM for maltose and of 0.16 muM for maltotroise was found. From the comparison of the kinetic parameters of transport of maltose and maltotriose in wild-type and lambda-resistant mutants with the binding constants for both sugars to purified maltose-binding protein, we conclude that the lambda receptor facilitates the diffusion of maltose and maltodextrins through the outer membrane.

Time dependent effects on contractile properties, fibre population, myosin light chains and enzymes of energy metabolism in intermittently and continuously stimulated fast twitch muscles of the rabbit.

Abstract: Fast-twitch tibialis anterior and extensor digitorum longus rabbit muscles were subjected to long-term intermittent (8 h daily) or continuous (24 h daily) indirect stimulation with a frequency pattern resembling that of a slow motoneuron. Increases in time to peak of isometric twitch contraction were observed without parallel changes in the pattern of myosin light chains or alterations in the distribution of slow and fast fibres as discernible by the histochemical ATPase reaction. However, changes in the fibre population and in the myosin light chain pattern were observed after intermittent stimulation periods exceeding 40 days or continuous stimulation periods longer than 20 days. Under these conditions even higher increases were found in contraction time. In one animal a complete change in fibre population was observed. In this case myosin light chains of the slow (LCS1, LCS2) and of the fast type (LCf1) were obviously synthetized simultaneously within the same fibre. Early changes in the enzyme activity pattern of energy metabolism indicated a conversion of the fibres including their mitochondrial population. These changes and the earlier reported changes in the sarcoplasmic reticulum are probably responsible for the early changes in contractile properties which occur before the transformation of the myosin.

Elastic effects of structural relaxation in glasses at low temperatures

Abstract: A complete theoretical description of the effect of the relaxation of two-state structural defects on the elastic properties of glasses at low temperatures is given. New experimental data on the ultrasonic attenuation in vitreous silica and borosilicate glass in the temperature range between 0.3 and 4 K and at frequencies between 30 and about 500 MHz are presented and explained by this theory. We propose that over the entire temperature range from our lowest temperature of 0.3 to about 100 K the attenuation is due to the relaxation of defects with similar atomic structure.

Polymorphism of a synthetic DNA in solution.

Abstract: EARLY work on the X-ray diffraction of DNA fibres showed that DNA can adopt different conformations, and up to now at least four different double-helical models have been described in detail: the A, B, C, and D forms1–4. During recent years, it has become likely that DNA in solution also occurs in different conformations5–7. For studying such helix–helix transitions synthetic polynucleotides with a defined sequence offer an important tool, since in natural DNA only a very small fraction of the molecule usually has a particular sequence and, sequence-specific effects of the conformation will therefore escape detection.

Kinetic analysis of carrier-mediated ion transport by the charge-pulse technique.

Abstract: The charge-pulse technique has been used previously for the study of quasistationary processes in membranes which required only a moderate time resolution. It is shown here that a time resolution of about 400 nsec may be achieved with this technique and that it may be applied to the kinetic analysis of carrier-mediated ion transport. By this method we have studied the transport of alkali ions through optically black monoolein membranes in the presence of the ion carrier valinomycin. All three relaxation processes that are predicted by theory have been resolved. From the relaxation times and the relaxation amplitudes the rate constants for the association (k R ) and the dissociation (k D ) of the ioncarrier complex, as well as the translocation rate constants of the complex (k MS ) and the free carrier (k S ) could be obtained. For 1m Rb+ at 25° C the values arek R =3×105m−1 sec−1,k D =2×105 sec−1,k MS =3×105 sec−1,k S =4×104 sec−1. The activation energies of the single rate constants which have been estimated from experiments at two different temperatures range between 50 and 90 kJ/mol.

Low-frequency Raman scattering by defects in glasses

Abstract: We describe the excess Raman intensity recently observed by Winterling in vitreous silica for frequency shifts $\ensuremath{\omega}l20$ ${\mathrm{cm}}^{\ensuremath{-}1}$ in terms of structural relaxation of defects, which can have two distinct states of polarizability differing by an amount $\ensuremath{\Delta}\ensuremath{\alpha}=0.6$ ${\mathrm{\AA{}}}^{3}$. All other properties of the defects (distribution of potential barriers) are taken from ultrasonic data. The correct temperature dependence is obtained. The experimentally observed value for the depolarization ratio in vitreous silica (0.30 \ifmmode\pm\else\textpm\fi{} 0.03) can be explained if the simplest possible form of the polarizability difference between the two states is assumed. We briefly discuss Raman scattering by quantum-mechanical tunneling states, which should be observable at very low temperatures.

Subsarcolemmal mitochondria and capillarization of soleus muscle fibers in young rats subjected to an endurance training. A morphometric study of semithin sections.

Abstract: Rats, 6 weeks old, were subjected to a program of endurance running for 3, 6 and 12 weeks 05 to 08 μm thick sections of Epon embedded soleus muscles were studied with morphometric methods In cross-sections the area occupied by subsarcolemmal mitochondria was independent of the age, but was 53% higher after 12 weeks of training The mean depth of the zones with subsarcolemmal mitochondria increased only 15% to about 09 μm Thus, the subsarcolemmal mitochondria showed a pronounced spreading at the muscle fiber surface in trained muscles — The number of capillaries per fiber decreased slightly in controls and increased not significantly in trained muscles It is concluded that the subsarcolemmal mitochondria supply the energy for the active transport of metabolites through the sarcolemma in oxidative muscle fibers, and that they are the limiting factor for endurance performance of the soleus muscle fibers because the changes in the capillarization were only small It is suggested that the subsarcolemmal and the interfibrillar mitochondria have different functions and may therefore represent different types of mitochondria which can be distinguished by their morphology as well as by their biochemical properties

Two Approaches To Estimating Public Expenditures

Abstract: Traditionally, the demand for public expenditures has been estimated using for explanatory variables the average values of per capita income, as well as other variables. The results of this approach are disappointing, due to the lack of a theoretical basis. The public choice model, on the other hand, uses the political decision-making process (median voter model) to explain expenditures. Taking the same set of data, it can be shown that the public choice approach yields superior results. It also offers a solution to the unfruitful discussion about the influence of "political" determinants of public expenditures

Pressure-Induced Valence Change in Cerium Phosphide

Abstract: The pressure-volume relationship for cerium monophosphide (CeP) has been determined to 200-kbar pressure. In this range the substance maintains the NaCl-type structure and exhibits abnormal compression. From the data we conclude that there is an electronic transition involving a change in the valence state of Ce from ${3}^{+}$ towards the ${4}^{+}$ state near 100-kbar pressure. Qualitative observations of the reflectivity of CeP under pressure support the valence transition. CeP under high pressure may be expected to exhibit superconductivity.

Insulin action on glucose transport and calcium fluxes in developing muscle cells in vitro.

Abstract: The glucose transport system of developing muscle cells in vitro is described. Uptake experiments showed stereospecificity but no saturation or competition. K,-values for early myoblasts could not be determined ; with progressive development, the absolute amount of uptake increases and becomes insulin sensitive. K,-values of about 13 mM basal and 32 mM for insulin stimulation were determined and discussed with respect to the glucose transport mechanism. Basal and insulindependent transport were shown to be dependent on the presence of calcium in the medium. Moreover, the ionophore A 23187 could be shown to mimic the insulin effect on glucose uptake. 45Ca saturation and desaturation experiments showed that insulin enhances the “mitochondrial” calcium pool by 35 and increases the apparent “cytoplasmic” efflux rate constant by 50 %. We propose that insulin increases the free cytoplasmic calcium concentration, which may be the intracellular signal for the stimulation of glucose transport.

Identification and Properties of 8‐Hydroxyflavin ‐Adenine Dinucleotide in Electron‐Transferring Flavoprotein from Peptostreptococcus elsdenii

Abstract: 1 A new flavin prosthetic group has been isolated in pure form from the electron-transferring flavoprotein of Peptostreptococcus elsdenii. Its structure has been established as the FAD derivative of 7-methyl-8-hydroxyisoalloxazine: Proof of this structure has been obtained by chemical synthesis of 7-methyl-8-hydroxyisoalloxazine models, and by stepwise degradation of the native compound to 7-methyl-8-hydroxyalloxazine. The orange chromophore is characterized by a strong absorption band with a maximum at 472 nm (ɛ= 41000 M−1 cm−1) and a pK at 4.8 due to the ionisation of the C(8)-OH group. 2 The properties of a series of functionally substituted, derivatives of 8-hydroxy flavins and lumichromes have been investigated to provide a basis for interpreting the effects of pH on the spectroscopic properties of the 8-hydroxy derivatives of FAD and FMN. 3 The 8-hydroxy derivative of FAD is bound by apo-d-amino acid oxidase; the complex shows no catalytic activity. The 8-hydroxy derivative of FMN is bound by apoflavodoxin to give a complex which has catalytic activity similar to that of native flavodoxin. The complex is reversibly reduced by dithionite, first to a relatively stable semiquinone and further to the dihydroflavin form.

Critical bands and filter characteristics in the ear of the housemouse (Mus musculus)

Abstract: Critical ratios (CRs) and critical bands (CBs, measured by narrow-band masking) were determined in one and the same test-procedure in 9 housemice (Mus musculus). Two spectrum levels of white band-pass noise (0 dB, 20 dB) and 9 frequencies between 2 kHz and 60 kHz were tested. CBs and CRs follow generally the same frequency-dependent functions. At 2 kHz the width of CRs is significantly larger (p 0.05) appear. No individual animal tested had significantly larger or smaller CBs (p>0.05) throughout the frequency range than the others. Critical bands represent equidistant parts of about 1 mm on the basilar membrane of the mouse. The steepness of the slopes of the CB-forming filter is directly proportional to frequency in the range from 10 kHz to 50 kHz. The critical band related distribution of behaviorally measurable excitation along the basilar membrane of the mouse is shown for the tested range of noise bands and spectrum levels. Present results are in essential agreement with respective data for man.

Agonistic behaviour and electric signalling in a mormyrid fish, Gnathonemus petersii

Abstract: 1. Agonistic motor behaviour and concurrent electric signalling were studied in individually held, residential Gnathonemus petersii. Aggressive behaviour was elicited by presenting a specimen of a closely related species, Mormyrus rume, for 3 min a day. 2. The principal agonistic motor patterns are described (Fig. 2). Among them head butt, approach and lateral display were further analysed. 3. The electrical activity displayed during agonistic behaviour was found to differ fundamentally both from isolated resting and swimming conditions. The mean discharge rate recorded during aggressive behaviour (31 Hz, Fig. 3 c) is approximately twice the rate observed in an isolated swimming fish (Fig. 3b) and three times the rate displayed by a resting animal (Fig. 3a). An attacking G. petersii exhibits a much greater range of electric organ discharge (EOD) intervals than isolated swimming or resting individuals. EOD-interval histograms recorded from attacking fish show two sharp modes at high discharge rate; there are no intermediate intervals. 4. During the course of an attack, the initially low and variable discharge rate increases fairly linearly as the distance from the attacked fish diminishes (Fig. 9). The EOD rate associated with physical contact (head butt) comprises between 60 and 80 Hz in 24 of 28 attacks analysed; the dominant mode of the distribution is 61 Hz (Fig. 8). 5. During subsequent lateral display, G. petersii emits a high discharge rate pattern consisting of two types of “steady-state” activities which may last up to a few seconds: the first is a fairly regular alternation of approx. 16 and 8 ms intervals (paired pulses); this pattern gives rise to the two peaks of high discharge rate in the interval histogram (Fig. 3c). The second is a regular sequence of either 16 or 8 ms intervals (Fig. 4A). The only female among the animals used in our study showed the same display but did not exhibit the highest possible discharge rate (i.e. a regular sequence or 8 ms intervals; Fig. 4B). The high discharge rate is terminated by a sudden discharge break (Figs. 4A and 6). 6. It is suggested that the attack-associated EOD rate increase is a remnant of an ordinary locomotory pattern which has changed its function to a ritualised aggressive signal that occurs in a socially significant and well-defined context. The high discharge rate might serve three functions: (i) behavioural isolation of closely related, sympatrically living mormyrids (perhaps by character displacement); (ii) recognition of sexes; (iii)_synchronisation of mates during courtship.

Synthesis, separation, identification and interconversion of riboflavin phosphates and their acetyl derivatives: a reinvestigation.

Abstract: A reinvestigation of flavin phosphate synthesis, separation, identification, and interconversion was made in view of contradictory results in the literature. It has been confirmed that monochlorophosphoric acid is the best agent for selective 5′-monophosphorylation of riboflavin and derivatives. This reaction yields, however, invariably up to 20% of an isomer, which has been separated by preparative thick-layer chromatography and shown to be the 4′-monophosphate. All the earlier authors failed to detect this isomer which does not bind to flavodoxin. It equilibrates in dilute mineral acid to yield an 8:2 mixture of 5′-phosphate to 4′-phosphate by phosphate migration. The formation of 2′,3′,4′-triacetyl-flavin mononucleotide, according to Christie, S. M. H., Kenner, G. W. & Todd, A. R. (1954) J. Chem. Soc., 46–52, upon acid-catalysed acetylation of pure FMN, was confirmed. The same reaction under base catalysis, however, does not yield 2′,3′,4′-triacetyl-flavin mononucleotide as claimed by Khomutova, E. D., Shapiro, T. A., Mezentseva, M. V. & Berezovskii, V. M. (1965) Otd. Ohshch. i Tekhn. Khim., 241–244, Chem. Absir. 65, 5516a, but in fact up to 80% 2′,3′-diacetyl-flavin 4′: 5′-cyclophosphate as the main product, which is stable under neutral and weak acidic conditions and does not hydrolyse to 2′,3′-diacetyl-flavin 5′-monophosphate as claimed by McCormick, D. B. (1974) J. Heterocycl. Chem. 11, 969–974. The various flavin phosphates and their acetyl derivatives have been identified by pH-titration, electrophoresis, and proton magnetic resonance spectrometry, which permit direct analyses of crude reaction products as well as a rapid purity check of commercial FMN.

The structure of the covalent flavin adduct formed between lactate oxidase and the suicide substrate 2-hydroxy-3-butynoate.

Abstract: Hydroxy-3-butynoic acid is a suicide substrate for Mycobacterium smegmatis lactate oxidase. Inactivation occurs by covalent modification of enzyme-bound FMN and does not involve labeling of the apoprotein. The spectrum of the enzyme bound adduct suggests that it is a 4a,5-dihydro- flavin derivative. When this adduct is released from the enzyme, a complex mixture of unstable compounds is obtained. When the initially formed enzyme-bound adduct is reduced with L-lactate oxidase (EC 1.13.12.4) from Mycobacterium smegmatis catalyzes the oxidative decarboxylation of L-lac- tate. OH I_

Kinetic studies on the inactivation of L-lactate oxidase by [the acetylenic suicide substrate] 2-hydroxy-3-butynoate.

Abstract: 2-Hydroxy-3-butynoate is both a substrate and an irreversible inactivator of the flavoenzyme L-lactate oxidase. The partitioning between catalytic oxidation of 2-hydroxy-3-butynoate and inactivation of the enzyme is determined by the concentration of the second substrate, O2. Rapid reaction studies show the formation of an intermediate which is common to both the oxidation and inactivation pathways. This intermediate appears to be a charge-transfer complex between enzyme-reduced flavin and 2-keto-3-butynoate. It is characterized by a long-wavelength absorbing band (gamma(max) 600 nm) and lack of fluorescence, making it easily distinguished from the subsequently formed inactivated enzyme, which has no long wavelength absorption (gamma(max) 318, 368 nm) and which is strongly fluorescent. Inactivation is also accomplished by reaction of the reduced enzyme with 2-keto-3-butynoate. The absorbance and fluorescence characteristics of the inactivated enzyme are similar to those of a model compound, C(4a), N(5)-propano-bridged FMN bound to apolactate oxidase. That the modified chromophore of the inactivated enzyme is an adduct involving both the C(4a) and N5 positions is further supported by the spectral and fluorescence changes resulting from treatment of the inactivated enzyme with borohydride.

Regulation of ion permeabilities of isolated rat liver cells by external calcium concentration and temperature.

Abstract: Regulation of ion transport through the plasma membrane was studied on single cell suspensions of hepatocytes, obtained after perfusion of rat liver with collagenase/hyaluronidase solution. Steady-state intracellular K and Na contents were shown to be markedly dependent on external Ca concentration and temperature, the sum of both ion concentrations remaining nearly constant. In contrast, steady-state intracellular chloride content was found to be independent of external Ca concentration, but dependent on temperature. Using the constant field relations, the passive permeabilitiesP K andP Cl for potassium and chloride, respectively, were derived from the experimental data. At temperatures at and above 37°C, with increasing external Ca concentration,P K exhibits a sharp decrease at about 10−4 m. In contrast,P Cl at 37°C was found to be independent of Ca concentration within experimental error. Earth alkali ions other than Ca, show marked but different effects onP K if compared at equal concentrations. Preincubation of the cells with cholesterol leads to a broadening of the dependence ofP K on external Ca concentration. The above results, as well as those on the dependence ofP K on external Ca concentration obtained by other authors, could be quantitatively described by a theoretical model of the plasma membrane proposed earlier. This model postulates regulatory binding sites, which cooperatively undergo a cation exchange of divalent cations by K+ ions from the external medium, if the cation composition of the latter is altered.

The effect of toxins from Ameonia sulcata (Coelenterata) on neuromuscular transmission and nerve action potentials in the crayfish ( Astacus leptodactylus )

Abstract: 1. The effects of three toxins (ATX I, II and III) from the sea anemoneAnemonia sulcata have been investigated on neuromuscular transmission and nerve action potentials in the crayfisch. 2. In the neuromuscular preparations the toxins produce a gradual increase in the amplitude of the excitatory junction potentials until they reach the threshold for the generation of electrically excitable membrane responses. This is followed by repetitive activity of the motor axons, first on single stimuli, later occurring spontaneously. 3. The effects of the three toxins differ only with regard to the effective concentrations: threshold concentrations amount to 5×10−9 M for ATX I and II, to 10−7 M for ATX III. 4. The toxins show no postsynaptic effects on the muscle fibre membrane and on their electrical responses upon direct stimulation. 5. All toxins prolong the action potentials of the giant axons in the abdominal nerve cord. The duration may reach more than one second. The rate of rise of the action potentials is not affected. In several cases, however, an increase of the amplitude of the action potential was observed. 6. The effects of ATX I and II are irreversible in contrast to those of ATX III. 7. It is suggested that the high potency of the toxins in the nanomolar range, their small size, the known composition and, in the case of ATX II, sequence of amino acids together with the specificity of action on the sodium channels, proved in different preparations, make these substances to important tools in the study of excitable membranes.