The influence of locust bean gum and dextran on the gelation of κ-carrageenan

The interaction of κ-carrageenan with locust bean gum and dextran has been studied by rheology, differential scanning calorimetry (DSC), and electron spin resonance spectroscopy (ESR). Rheological measurements show that the carrageenan gel characteristics are greatly enhanced in the presence of locust bean gum but not in the presence of dextran. Carrageenan/locust bean gum mixtures show two peaks in the dsc cooling curves. The higher temperature peak corresponds to the temperature of gelation and its intensity increases at the expense of the lower temperature peak as the proportion of locust bean gum in the mixture increases. Furthermore, the DSC heating curves show enhanced broadening when locust bean gum is present, indicating increased aggregation. These results are taken as evidence of carrageenan/locust bean gum association. The gelation process has also been followed by ESR using spin-labeled carrageenan. On cooling carrageenan solutions, an immobile component appears in the ESR spectra signifying a loss of segmental mobility consistent with chain stiffening due to the coil → helix conformational transition and helix aggregation. For carrageenan/locust bean gum mixtures, carrageenan ordering occurs at temperatures corresponding to the higher temperature DSC setting peak and the temperature of gelation. Similar studies using spin-labeled locust bean gum show that its mobility remains virtually unaffected during the gelation process. It is evident, therefore, that carrageenan and locust bean gum interact only weakly. It is proposed that at low carrageenan concentrations the gel network consists of carrageenan helices cross-linked by locust bean gum chains. At high carrageenan concentrations the network is enhanced by the additional self-aggregation of the “excess” carrageenan molecules. For carrageenan/dextran mixtures, only one peak is observed in the dsc cooling curves. The onset of gelation shifts to higher temperatures only at very high (20%) dextran concentrations and this is attributed to volume exclusion effects. Furthermore, there is no enhanced broadening of the peaks in the DSC heating curves as for the carrageenan/locust bean gum systems. It is therefore concluded that carrageenan/dextran association does not occur. The difference in behavior between locust bean gum and dextran is attributed to the greater flexibility of the dextran chains. © 1996 John Wiley & Sons, Inc.     

A new bacterial polysaccharide (YAS34). I. Characterization of the conformations and conformational transition

This paper concerns the study of the conformational transition of a new exopolysaccharide (YAS34) using experimental techniques such as optical rotation, conductimetric and microcalorimetric measurements as a function of temperature. The behaviors of this polysaccharide in the acid or sodium salt form are compared; a deacetylated sample is also prepared to demonstrate the role of substituents. For the native structure (never heated), a conformational transition is observed but the deacetylated polysaccharide exhibits no ordered conformation. Multidetection size exclusion chromatography (SEC) analyses and conductimetric experiments allowed to determine the nature of each conformation and the molecular dimensions. From these results, it is suggested that the native conformation is a double helix which by heating over T(m) (temperature corresponding to half conformational transition) dissociates into disordered single chains. In the acid and sodium salt forms, by cooling below T(m), an ordered conformation is restored. This conformation seems to be an intramolecular double helix 'hairpin-like turn' (called renatured conformation). Nevertheless an irreversible denaturation is obtained progressively in the sodium salt form when the time of heating over T(m) increases. The conformation of the deacetylated polysaccharide corresponds to that of a single flexible chain (disordered conformation). The conformational transition for the native conformation was studied also in relation to the polyelectrolytic character of the polysaccharide: stability as a function of salt nature and salt and polymer concentrations was investigated for the polymer initially in the sodium and acid forms.     

Cation effects on sol-gel and gel-sol phase transitions of kappa-carrageenan-water system

Sol–gel and gel–sol phase transitions of κ-carrageenan in pure water and in KCl solution were studied using photon transmission technique. Photon transmission intensity, I_tr, was monitored against temperature to determine the sol–gel and gel–sol temperatures (T_sg and T_gs) and activation energies (ΔH_sg and ΔH_gs). It was observed that T_gs was notably higher than T_sg due to the hysteresis on the phase transition loops. T_gs and ΔH_gs values were also higher for gels containing KCl than for those without KCl. The increase in carrageenan content caused an increase in both critical temperatures and activation energies for the gels prepared in pure water and in KCl solution. Increases in the KCl/carrageenan ratio, raised both T_gs and T_sg. Similarly ΔH_sg was elevated by the increase in cation content of the gel. These results were interpreted as the formation of stronger gels in the presence of KCl in water.     

Kinetics for exchange of imino protons in the d(C-G-C-G-A-A-T-T-C-G-C-G) double helix and in two similar helices that contain a G . T base pair, d(C-G-T-G-A-A-T-T-C-G-C-G), and an extra adenine, d(C-G-C-A-G-A-A-T-T-C-G-C-G)

The relaxation lifetimes of imino protons from individual base pairs were measured in (I) a perfect helix, d(C-G-C-G-A-A-T-T-C-G-C-G), (II) this helix with a G . C base pair replaced with a G . T base pair, d(C-G-T-G-A-A-T-T-C-G-C-G), and (III) the perfect helix with an extra adenine base in a mismatch, d(C-G-C-A-G-A-A-T-T-C-G-C-G). The lifetimes were measured by saturation recovery proton nuclear magnetic resonance experiments performed on the imino protons of these duplexes. The measured lifetimes of the imino protons were shown to correspond to chemical exchange lifetimes at higher temperatures and spin-lattice relaxation times at lower temperatures. Comparison of the lifetimes in these duplexes showed that the destabilizing effect of the G . T base pair in II affected the opening rate of only the nearest-neighbor base pairs. For helix III, the extra adenine affected the opening rates of all the base pairs in the helix and thus was a larger perturbation for opening of the base pairs than the G . T base pair. The temperature dependence of the exchange rates of the imino proton in the perfect helix gives values of 14-15 kcal/mol for activation energies of A . T imino protons. These relaxation rates were shown to correspond to exchange involving individual base pair opening in this helix, which means that one base-paired imino proton can exchange independent of the others. For the other two helices that contain perturbations, much larger activation energies for exchange of the imino protons were found, indicating that a cooperative transition involving exchange of at least several base pairs was the exchange mechanism of the imino protons. The effects of a perturbation in a helix on the exchange rates and the mechanisms for exchange of imino protons from oligonucleotide helices are discussed.     

Mechanical stability of single DNA molecules

Using a modified atomic force microscope (AFM), individual double-stranded (ds) DNA molecules attached to an AFM tip and a gold surface were overstretched, and the mechanical stability of the DNA double helix was investigated. In lambda-phage DNA the previously reported B-S transition at 65 piconewtons (pN) is followed by a second conformational transition, during which the DNA double helix melts into two single strands. Unlike the B-S transition, the melting transition exhibits a pronounced force-loading-rate dependence and a marked hysteresis, characteristic of a nonequilibrium conformational transition. The kinetics of force-induced melting of the double helix, its reannealing kinetics, as well as the influence of ionic strength, temperature, and DNA sequence on the mechanical stability of the double helix were investigated. As expected, the DNA double helix is considerably destabilized under low salt buffer conditions (相似文献   

The kinetics and thermodynamics of reversible denaturation of crystalline soybean trypsin inhibitor

Crystalline soybean trypsin inhibitor protein undergoes denaturation on heating which is reversed on cooling. In the range of temperature of 35 to 50 degrees C. a solution of the protein consists of a mixture of native and denatured forms in equilibrium with each other. The equilibrium is only slowly established and its final value at any temperature is the same whether a heated, denatured solution of the protein is cooled to the given temperature or whether a fresh solution is raised to that temperature. The kinetics of reversible denaturation of the soybean protein as well as the reversal of denaturation is that of a reversible unimolecular reaction, each process consisting at a given temperature of the same two simultaneous reactions acting in opposite directions. The experimental data on the effect of temperature on the velocity and the equilibrium constants of the opposing reaction were utilized in evaluating the reaction energies and activation energies. The reaction energies for denaturation were found to be as follows:- Change in total heat of reaction DeltaH = 57,000 calories per mole Change in entropy of reaction DeltaS = 180 calories per degree per mole The heat of activation DeltaH(1) (double dagger) for denaturation = 55,000 The heat of activation DeltaH(2) (double dagger) for the reversal of denaturation = -1900 The entropy DeltaS(1) (double dagger) for denaturation = 95 The entropy DeltaS(2) (double dagger) for reversal of denaturation = -84     

Gel-forming structures and stages of red algal galactans of different sulfation levels

Sol-gel transition processes of algal galactans were studied using cryofixation method in combination with freeze-drying and scanning electron microscopy (SEM) techniques. The structures formed in successive stages of gelling process upon cooling were rapidly frozen at defined temperature points and viewed by SEM. It was established that in the case of both types of gelling galactans investigated, a fine honeycomb-like network exists for a wide range of solution temperatures. The formation and structure of this network depends on the structural type, gelling stage, and concentration of the galactan in solution. The honeycomb suprastructures exist also in carrageenan and agarose sols (at temperatures considerably exceeding the gelling temperatures). An additional helical network formed showed different behaviour in the case of carrageenan and agar-type polysaccharides. In the gel-formation process, tightening of the network takes place in both types of galactan gels; the honeycomb structures persist in carrageenan (furcellaran) but not in agarose gels.     

Rheology of concentrated isotropic and anisotropic xanthan solutions: 3. Temperature dependence

The oscillatory rheology of one rodlike and one semiflexible xanthan sample has been investigated as a function of temperature in the range of xanthan concentrations where the polymer forms a lyotropic liquid crystalline phase in aqueous NaCl solutions. Readily observed changes in the rheological observables at temperatures corresponding to phase boundaries permit construction of the biphasic chimney region of the temperature-composition phase diagram. The chimney region leans toward larger values of the polymer concentration with increasing temperature, presumably as a consequence of a reduction in the effective axial ratio of the helical polymer with increasing temperature. The results permit construction of plots of the rheological observables as a function of polymer concentration at temperatures T in the range 20 相似文献   

Determination of L(alpha)-H(II) phase transition temperature for 1,2-dioleoyl-sn-glycero-3-phosphatidylethanolamine

The thermodynamic properties of fully-hydrated lipids provide important information about the stability of membranes and the energetic interactions of lipid bilayers with membrane proteins (Nagle and Scott, Physics Today, 2:39, 1978). The lamellar/inverse hexagonal (L(alpha)-H(II)) phase transition of 1,2-dioleoyl-sn-glycero-3-phosphatidylethanolamine (DOPE) water mixtures is a first-order transition and, therefore, at constant pressure, must have a thermodynamically well-defined equilibrium transition temperature. The observed transition temperature is known to be dependent upon the rate at which the temperature is changed, which accounts for the many different values in the literature. X-ray diffraction was used to study the phase transition of fully-hydrated DOPE to determine the rate-independent transition temperature, T(LH). Samples were heated or cooled for a range of rates, 0.212 < r < 225 degrees C/hr, and the rate-dependent apparent phase transition temperatures, T(A)(r) were determined from the x-ray data. By use of a model-free extrapolation method, the transition temperature was found to be T(LH) = 3.33 +/- 0.16 degrees C. The hysteresis, /T(A)(r) - T(LH)/, was identical for heating and cooling rates, +/-r, and varied as /r/beta for beta approximately 1/4. This unexpected power-law relationship is consistent with a previous study (Tate et al., Biochemistry, 31:1081-1092, 1992) but differs markedly from the exponential behavior of activation barrier kinetics. The methods used in this study are general and provide a simple way to determine the true mesomorphic phase transition temperatures of other lipid and lyotropic systems.     

Temperature dependence and Arrhenius activation energy of F-actin velocity generated in vitro by skeletal myosin.

The effect of temperature on the velocity of rhodamine phalloidin-labelled F-actin moving in vitro on rabbit skeletal myosin has been studied. Translating actin filaments were visualized by epi-fluorescence in an inverted microscope, equipped with temperature control (+/- 0.2 K) of the stage and objective. Images were recorded in real time at magnifications of 400x or 160x by an intensified CCD camera on video tape. Motion of individual filaments was tracked by hand and velocities determined using frame times recorded simultaneously on the video tape. Velocity changed from 12 microns per second at 42 degrees C to 11 nm per second at 3 degrees C. The Arrhenius plot is non-linear, with the data following a cubic regression curve with no evident breaks or jumps. Data taken over the temperature range from single preparations followed the same curve for both heating and cooling; this indicates reversibility and absence of hysteresis. A hyperbolic model that smoothly translates with temperature between two asymptotic activation energies fits the data above 7 degrees C: these energies are 50(+/- 5) kJ per mole (Q10 = 1.9) at high temperatures and 289(+/- 29) kJ per mole (Q10 = 76.5) at low temperature with a transition temperature of 15.4(+/- 0.6) degrees C. These values are compared with other measurements made in vitro, in solution studies and on muscle fibres. An Arrhenius activation energy of 50 kJ per mole and a transition temperature of 15 degrees C are consistent with previous determinations but 289 kJ per mole is significantly greater than has been seen at low temperatures in other systems. This may indicate a different rate-limiting step in the kinetics of skeletal myosin driving actin filaments in vitro below 15 degrees C. Current determinations of the myosin "step-size" assume that the actin velocity is determined by the rate of ATP hydrolysis; the data confirm similar activation energies above 20 degrees C but they show that the temperature dependencies and activation energies are different at lower temperatures, implying uncoupling of the two processes.     

Binding effect of Cu(2+) as a trigger on the sol-to-gel and the coil-to-helix transition processes of polysaccharide, gellan gum

The binding effect of divalent cation Cu(2+) on the gelation process with a coil-helix transition in Cu(2+)/gellan aqueous solutions has been successfully elucidated by EPR, CD, and viscoelasticity measurements. Generally, Na-type gellan gum in aqueous solution can make gel when accompanied by an intrinsic coil-helix formation induced by hydrogen bonding between chains without any additional cations at T(ch)(-)(in) ( approximately 29 degrees C) with cooling temperature. An extrinsic coil-helix transition, induced by additional divalent cations in advance of the intrinsic sol-gel transition of gellan gum, is separately detected by CD measurement. The extrinsic coil-helix transition temperatures T(ch)(-)(ex) (>47 degrees C), which increased with the Cu(2+) concentration added, were nearly identical to the sol-gel transition temperature, T(sg), determined by the viscoelasticity measurement. Judging from the molar ellipticity by CD measurement and quantitative analysis of EPR spectra, it was elucidated that the helix forming process via divalent cations is composed of two steps ascribed to the different origins, i.e., a chemical binding effect via Cu(2+) ions in the initial stage and hydrogen bonds subsequently. Finally, we propose the coil-helix and the sol-gel transition mechanism initiated by the binding effect with the divalent cation, in which the partial chelate formation can cause local formation of helices and junction zones in the vicinity of the chelates at the initial stage of the process and stabilize the helices and the junction zones. On the other hand, the stabilized helices and junction zones can induce further formation and further stabilization of the Cu(2+)-gellan chelates. The mutual stabilization promotes the formation of three-dimensional network structure at the higher temperature than the intrinsic temperature for network formation.     

Calorimetric and chiroptical evidence of aggregate-driven helix formation in carrageenan systems

Thermally induced, order-disorder transitions of iota- and kappa-carrageenan have been monitored by optical rotation and differential-scanning calorimetry in various ionic environments. Conformational ordering in kappa-carrageenan is observed only in the presence of cations that have been shown previously to promote helix-helix aggregation, and shows marked hysteresis between heating and cooling. Iota-carrageenan, by contrast, shows an order-disorder transition in the non-aggregating, tetramethylammonium salt form, at substantially lower temperature than for kappa-carrageenan, and without hysteresis. In the presence of potassium ions, which are known to promote aggregation, iota-carrageenan shows two distinct thermal-transitions, one without hysteresis at the same temperature as observed under non-aggregating conditions, and one with significant hysteresis close to the temperature of the kappa-carrageenan transition. We interpret these transitions as helix-to-coil and aggregated helix-to-coil, respectively. This interpretation is supported by measurements of the enthalpy changes of the transitions; ΔH values show a systematic increase with increasing aggregation and hysteresis. We conclude that the double helix of iota-carrageenan can exist as a stable entity in isolation, but may be further stabilised by aggregation, whereas the kappa-carrageenan helix is stable only when aggregated.     

Effect of temperature and pH on the β–helix transition of poly(L-lysine) in sodium dodecyl sulfate solution

The conformational phase diagram of poly(L -lysine) (4.6 × 10^?4 M, residue) in sodium dodecyl sulfate (1.6 × 10^?2 M) solution was constructed from circular dichroism results at various temperatures and pH's. Poly(L -lysine)–sodium dodecyl sulfate complexes undergo a β–helix transition upon raising the pH of the solution. The transition pH tends to shift downward at elevated temperatures. No helix–β transition can be detected for poly(L -lysine) in sodium dodecyl sulfate solution (pH > 11) even after 1-hr heating at 70°C. This is in marked contrast with uncharged poly(L -lysine) solution without sodium dodecyl sulfate, which is converted into the β-form upon mild heating of the solution above 50°C.     

Kinetic phase behavior of distearoylphosphatidylethanolamine dispersed in glycerol

Phase behavior of distearoylphosphatidylethanolamine dispersed in excess glycerol has been examined by differential scanning calorimetry. Transformation from lamellar-gel to lamellar crystalline phase was found to take place at temperatures near 74.9 degrees C upon cooling and near 76.3 degrees C during heating scans. The transition can also be observed under isothermal conditions at temperature in this range. The kinetics of the transformation from lamellar-gel to lamellar-crystal phase was analyzed by the well-known Avrami equation. The apparent Avrami exponents were found to be approximately 1.6. The effective dimensionality of the growth pattern can then be set as 1, after taking into account the contribution of nucleation at the examination temperatures. The activation energy of the phase transition was estimated as approximately 255 kJ mol(-1). The data are discussed in terms of development of successful cryoprotective strategies using glycerol.     

Thermotropic phase behavior of model membranes composed of phosphatidylcholines containing cis-monounsaturated acyl chain homologues of oleic acid: differential scanning calorimetric and 31P NMR spectroscopic studies

The thermotropic phase behavior of dioleoylphosphatidylcholine and six of its longer chain homologues was studied by differential scanning calorimetry and 31P nuclear magnetic resonance (NMR) spectroscopy. Aqueous dispersions of these compounds all exhibit a single endotherm upon heating but upon cooling exhibit at least two exotherms, both of which occur at temperatures lower than those of their heating endotherm. The single transition observed upon heating was shown by 31P NMR spectroscopy to be a net conversion from a condensed, subgel-like phase (Lc phase) to the liquid-crystalline state. Aqueous ethylene glycol dispersions of these compounds also exhibit single endotherms upon heating and cooling exotherms centered at temperatures lower than those of their corresponding heating endotherm. However, the behavior of the aqueous ethylene glycol dispersions differs with respect to their transition temperatures and enthalpies as well as the extent of "undercooling" observed, and there is some evidence of discontinuities in the cooling behavior of the odd- and even-numbered members of the homologous series. Like the aqueous dispersions, 31P NMR spectroscopy also shows that the calorimetric events observed in aqueous ethylene glycol involve net interconversions between an Lc-like phase and the liquid-crystalline state. However, the Lc phase formed in aqueous ethylene glycol dispersions exhibits a considerably broader powder pattern than that observed in water. This, together with the fact that the transition enthalpies of the aqueous ethylene glycol dispersions are considerably higher than those of the aqueous dispersions, indicates that these lipids form more ordered Lc phases in aqueous ethylene glycol.(ABSTRACT TRUNCATED AT 250 WORDS)     

Kinetic Hysteresis in Collagen Folding

The triple helix of collagen shows a steep unfolding transition upon heating, whereas less steep and more gradual refolding is observed upon cooling. The shape of the hysteresis loop depends on the rate of temperature change as well as the peptide concentration. Experimental heating and cooling rates are usually much faster than rates of unfolding and refolding. In this work, collagen model peptides were used to study hysteresis quantitatively. Their unfolding and refolding profiles were recorded at different heating and cooling rates, and at different peptide concentrations. Data were fitted assuming kinetic mechanisms in which three chains combine to a helix with or without an intermediate that acts as a nucleus. A quantitative fit was achieved with the same kinetic model for the forward and backward reactions. Transitions of exogenously trimerized collagen models were also analyzed with a simplified kinetic mechanism. It follows that true equilibrium transitions can only be measured at high concentrations of polypeptide chains with slow scanning rates, for example, 0.1°C/h at 0.25 mM peptide concentration of (Gly-Pro-Pro)₁₀. (Gly-Pro-4(R)Hyp)₁₀ folds ∼2000 times faster than (Gly-Pro-Pro)₁₀. This was explained by a more stable nucleus, whereas the rate of propagation was almost equal. The analysis presented here can be used to derive kinetic and thermodynamic data for collagenous and other systems with kinetically controlled hysteresis.     

Tobacco mosaic virus protein: kinetic and equilibrium studies on the pH-dependent transition. A-protein yields double disc.

Concentration dependent and temperature dependent stopped-flow experiments on the transition A-protein→double disc show a triphasic reaction consisting of a nucleation phase, a propagation phase and a slow redistribution of polymer size which involves the dissociation of “overshoot” aggregates into double discs and smaller aggregates. No first order rate process is observed under the present experimental conditions. Equilibrium circular dichroism data and preliminary kinetic data at various temperatures indicate a change at about 21°C which might be correlated to a partial transition double disc→helix parallelled by a further shift in the equilibrium of double disc formation; from both data the thermodynamic and activation parameters for the A-protein→double disc transition are estimated.     

The effect of the phase transition on the hydration and electrical conductivity of phospholipids.

Adsorption isotherms for various saturated phosphatidylcholines have been obtained. Lipids above and below their phase transition temperature differ only in the amount of water adsorbed and not in the nature of their adsorption isotherms. Cholesterol has an effect similar to that of increasing unsaturation in the hydrocarbon chains. Decreasing the length of the hydrocarbon chains for lipids below their phase transition temperature has no effect on the isotherms. If the chain length is short enough so that the lipids are above their transition temperature, however, a large increase in water adsorption occurs. All of the phospholipids exhibit a rapid increase of electrical conductivity for a few water molecules adsorbed per lipid molecule. All of the phospholipids show a saturation in conductivity at greater amounts of adsorbed water; the shape of the saturation region depends on whether the lipids are above or below their phase transition temperature. The activation energy for the electrical conductivity process depends on whether the hydrated lipids are in the "liquid-like" of the crystalline state, being lower for phospholipids in the liquid-like state. If the lipids are hydrated above their phase transition temperatures, their activation energies are lower than if they are hydrated below the transition temperature. Cholesterol lowers the activation energy. The phosphatidylcholines can be characterized by different activation energies, depending both upon their physical state and the presence of unsaturation in their hydrocarbon chains.     

Transition state characterization of the low- to physiological-temperature nondenaturational conformational change in bovine adenosine deaminase by slow scan rate differential scanning calorimetry

Bovine adenosine deaminase undergoes a nondenaturational conformational change at 29 degrees C upon heating which is characterized by a large increase in heat capacity. We have determined the transition state thermodynamics of the conformational change using a novel application of differential scanning calorimetry (DSC) which employs very slow scan rates. DSC scans at the conventional, and arbitrary, scan rate of 1 degree C/min show no evidence of the transition. Scan rates from 0.030 to 0.20 degrees C/min reveal the transition indicating it is under kinetic control. The transition temperature T(t) and the transition temperature interval deltaT increase with scan rate. A first order rate constant k1 is calculated at each T(t) from k1 = r(scan)/deltaT, where r(scan) is the scan rate, and an Arrhenius plot is constructed. Standard transition state analysis reveals an activation free energy deltaG(double dagger) of 88.1 kJ/mole and suggests that the conformational change has an unfolding quality that appears to be on the direct path to the physiological-temperature conformer.     

A fluorescence study on the gel-to-sol transition of kappa-carrageenan

The fluorescence technique was employed to study gel-to-sol transitions in kappa-carrageenan systems with various carrageenan contents. Pyranine was used as a fluorescence probe for monitoring these transitions. Fluorescence, Ip and scattered light, Isc intensities were measured against temperature to determine critical phase transition temperatures and exponents. It was observed that gel-to-sol transition temperatures, Tgs are found to be dependent to the carrageenan content. The measured critical exponents gamma' and beta' were found to be very close to the weight average degree of polymerization, DPw and gel fraction G, exponents (gamma and beta) of the classical Percolation Model.