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PubMed2006

Interaction of poly(L-lysines) with negatively charged membranes: an FT-IR and DSC study.

The influence of the binding of poly(L-lysine) (PLL) to negatively charged membranes containing phosphatidylglycerols (PG) was studied by DSC and FT-IR spectroscopy. We found a general increase in the main transition temperature as well as increase in hydrophobic order of the membrane upon PLL binding. Furthermore we observed stronger binding of hydration water to the lipid head groups after PLL binding. The secondary structure of the PLL after binding was studied by FT-IR spectroscopy. We found that PLL binds in an alpha-helical conformation to negatively charged DPPG membranes or membranes with DPPG-rich domains. Moreover we proved that PLL binding induces domain formation in the gel state of mixed DPPC/DPPG or DMPC/DPPG membranes as well as lipid remixing in the liquid-crystalline state. We studied these effects as a function of PLL chain length and found a significant dependence of the secondary structure, phase transition temperature and domain formation capacity on PLL chain length and also a correlation between the peptide secondary structure and the phase transition temperature of the membrane. We present a system in which the membrane phase transition triggers a highly cooperative secondary structure transition of the membrane-bound peptide from alpha-helix to random coil.

Binding Sites↗

PubMed1985

Cross polarization P-31 nuclear magnetic resonance of phospholipids.

P-31 single-pulse and cross-polarization (CP) nuclear magnetic resonance spectra were obtained of aqueous dispersions of pure phospholipids. Dimyristoyl phosphatidylcholine, dipalmitoylphosphatidylcholine, 1-palmitoyl-2-oleoyl phosphatidylcholine, egg phosphatidylcholine, bovine brain sphingomyelin, and transphosphatidylated (from egg phosphatidylcholine) phosphatidylethanolamine were studied. The spectra from all the phospholipids, taken in the usual single-pulse mode, showed the pseudo-axially symmetric powder pattern typical of phospholipids in a hydrated lamellar form. P-31 CP spectra of all the phosphatidylcholines and phosphatidylethanolamine revealed a decrease in intensity in the vicinity of the isotropic chemical shift as long as the lipid was above the gel-to-liquid crystalline phase transition temperature. This intensity pattern has been observed previously for C-13 CP spectra of molecules rotating rapidly about a single well-defined axis (e.g., solid benzene) (Pines, A., M.G. Gibby, and J.S. Waugh, 1973, J. Chem. Phys., 59:569-590). Pure lipid dispersions below their gel-to-liquid crystalline phase transition temperature, including dipalmitoylphosphatidylcholine and sphingomyelin, do not exhibit a local minimum in the CP spectrum at the position of the isotropic chemical shift. Thus, below the phase transition temperature, there is not the same rapid rotation of the headgroup about a well-defined axis. A dramatic change in the rate of headgroup rotation is shown to take place at the pretransition of dipalmitoylphosphatidylcholine. P-31 CP spectra were also obtained for bovine rod outer segment disk membranes, rabbit muscle sarcoplasmic reticulum membranes, a total lipid extract of the latter, and a recombined membrane containing human erythrocyte glycophorin. The CP spectra were similar to the single-pulse spectra, indicating a substantial difference in behavior from pure phospholipid dispersions. This is interpreted in terms of a slower headgroup rotation.

Animals↗

PubMed2005

Simulation of dynamical properties of normal and superfluid helium.

The formation of a superfluid when (4)He is cooled below the characteristic lambda transition temperature is accompanied by intricate quantum mechanical phenomena, including the emergence of a Bose condensate. A combination of path integral and semiclassical techniques is used to calculate the single-particle velocity autocorrelation function across the normal-to-superfluid transition. We find that the inclusion of particle exchange alters qualitatively the shape of the correlation function below the characteristic transition temperature but has no noticeable effect on the dynamics in the normal phase. The incoherent structure factor extracted from the velocity autocorrelation function is in very good agreement with neutron scattering data, reproducing the width, height, frequency shift, and asymmetry of the curves, as well as the observed increase in peak height characteristic of the superfluid phase. Our simulation demonstrates that the peak enhancement observed in the neutron scattering experiments below the transition temperature arises exclusively from particle exchange, illuminating the role of Bose-statistical effects on the dynamics of the quantum liquid.

Journal Article↗

PubMed1976

Role of phospholipids in the calcium-dependent ATPase of the sarcoplasmic reticulum. Enzymatic and ESR studies with phospholipid-replaced membranes.

Three types of partially purified ATPase enzymes having different phospholipid contents and compositions have been prepared: (a) an enzyme whose phospholipid moiety has been replaced predominantly by dioleoyl lecithin (DOL-enzyme), with about the same phospholipid content as the original sarcoplasmic reticulum, (b) dipalmitoyl lecithin-replaced enzyme whose phospholipid content is 30% of that of DOL-enzyme (DPL-enzyme), and (c) a partially delipidated enzyme with about the same phospholipid content as DPL-enzyme but with the original sarcoplasmic reticulum phospholipid composition (del-enzyme). The temperature dependence of Ca2+-activated ATPase activity of these preparations showed clearcut differences; with DOL-enzyme there was no appreciable break in the Arrhenius plot in the 3-40 degrees range; DPL-enzyme showed a break at 29 degrees, and del-enzyme and sarcoplasmic reticulum one at 18 degrees. Transition temperatures obtained from ESR studies with the use of spin-labeled stearic acid incorporated into the membranes agreed with those derived from ATPase assays. Thermo-dynamic analysis of the ATP hydrolysis rates shows that DPL-enzyme has considerably larger values of activation enthalpy and activation entropy below the transition temperature (29 degrees) than those of the other preparations, while all enzyme preparations show similar free energies of activation. The ESR data show that below their transition temperatures DPL-enzyme, and to a lesser degree del-enzyme, have a strongly restricted motion of their phospholipid molecules as compared with either DOL-enzyme or sarcoplasmic reticulum. Studies on the formation and decomposition of phosphoenzyme have been carried out with the three types of ATPase preparations. At 0 degrees, the rate of inorganic phosphate liberation is 8 times lower in DPL-enzyme than in del-enzyme with little difference in the steady state level of phosphoenzyme. In DOL-enzyme, the level of phosphoenzyme and the rate of inorganic phosphate liberation are 1.8 and 3.5 times higher than the corresponding values obtained with del-enzyme. Addition of ADP to the phosphorylated intermediate of DPL-enzyme induces a fast reversal of the phosphorylation reaction. These results indicate that the physical state of the phospholipid molecules associated with the enzyme affects the decomposition of phosphoenzyme, with little effect on the phosphorylation reaction and its reversal.

Adenosine Triphosphatases↗

PubMed1979

Fusion of phosphatidylserine and mixed phosphatidylserine-phosphatidylcholine vesicles. Dependence on calcium concentration and temperature.

Dynamic light scattering has been used to study the temperature dependence of Ca2+-induced fusion of phosphatidylserine vesicles and mixed vesicles containing phosphatidylserine and different phosphatidylcholines. The final vesicle size after Ca2+ and EDTA incubation serves as a measure of the extent of fusion. With phosphatidylserine vesicles, the extent of fusion shows a sharp maximum at an incubation temperature which depends on the Ca2+ concentration between 0.8 and 2 mM. The shift in the fusion peak temperature with Ca2+ concentration is similar to the typical shift in the phase transition temperature with divalent cation concentration in acidic phospholipids. The results suggest a direct correlation between the fusion peak temperature and the phase transition temperature in the presence of Ca2+ prior to fusion. With mixed vesicles containing up to 33% of a phosphatidylcholine in at least 2 mM Ca2+, the extent of fusion as a function of incubation temperature also shows a maximum. The fusion peak temperature is essentially independent of the quantity and type of phosphatidylcholine and the Ca2+ concentration, and identical to that with pure phosphatidylserine in excess Ca2+. The results imply that Ca2+- induced molecular segregation occurs first, and fusion subsequently takes place between pure phosphatidylserine domains.

Calcium↗

PubMed1981

Phase transitions in lipid bilayers. A theoretical model for phosphatidylethanolamine and phosphatidic acid bilayers.

We present an application of an earlier model to the description of phase transitions in phosphatidylethanolamine and phosphatidic acid bilayers. For the former system we modify the model to allow for very slightly closer chain packing. This small modification alone is sufficient to shift the main phase transition temperature to higher values, and to eliminate the lower transition. For the charged bilayers we incorporate electrostatic contributions into the free energy using double layer theory and we consider the excluded volume of counterions at the bilayer/water interface. Phase transition temperatures and the presence of absence of a lower transition are determined for various pH values, and the results are compared with experiment.

Hydrogen-Ion Concentration↗

PubMed2006

Freezing-in and production of entropy in vitrification.

Following the classical concepts developed by Simon [Z. Anorg. Allg. Chem. 203, 219 (1931)], vitrification in the cooling of glass-forming melts is commonly interpreted as the transformation of a thermodynamically (meta)stable equilibrium system into a frozen-in, thermodynamically nonequilibrium system, the glass. Hereby it is assumed that the transformation takes place at some well-defined sharp temperature, the glass transition temperature Tg. However, a more detailed experimental and theoretical analysis shows that the transition to a glass proceeds in a broader temperature range, where the characteristic times of change of temperature, tauT=-(TT), and relaxation times, tau, of the system to the respective equilibrium states are of similar order of magnitude. In this transition interval, the interplay of relaxation and change of external control parameters determines the value of the structural order parameters. In addition, irreversible processes take place in the transition interval, resulting both in an entropy freezing-in as well as in an irreversible increase of entropy and, as a result, in significant changes of all other thermodynamic parameters of the vitrifying systems. The effect of entropy production on glass transition and on the properties of glasses is analyzed here for the first time. In this analysis, the structural order-parameter concept as developed by de Donder and van Rysselberghe [Thermodynamic Theory of Affinity (Stanford University Press, Stanford, 1936)] and Prigogine and Defay [Chemical Thermodynamics (Longmans, London, 1954)] is employed. In the framework of this approach we obtain general expressions for the thermodynamic properties of vitrifying systems such as heat capacity, enthalpy, entropy, and Gibbs' free energy, and for the entropy production. As one of the general conclusions we show that entropy production has a single maximum upon cooling and two maxima upon heating in the glass transition interval. The theoretical concepts developed allow us to explain in addition to the thermodynamic parameters also specific features of the kinetic parameters of glass-forming melts such as the viscosity. Experimental results are presented which confirm the theoretical conclusions. Further experiments are suggested, allowing one to test several additional predictions of the theory.

Journal Article↗

PubMed1978

Physiological conditions affecting Staphylococcus aureus susceptibility to staphylococcin 1580.

Loss of salt tolerance, irreversible loss of viability, inhibition of l-glutamic acid uptake and effects on the high energy state of the membrane were used as parameters to measure the injury induced by staphylococcin 1580, a bacteriocin of Staphylococcus epidermidis, in susceptible cells of Staphylococcus aureus Oxford 209P. A small part of a growing cell population appeared to be temporarily resistant to the bacteriocin, and the cells were arrested in this stage when suspended in buffer. The proportion of susceptible cells may rapidly shift during exponential growth, apparently concomitantly with a change in cell metabolism. Glucose- and pyruvate-grown cells were equally susceptible to salts after staphylococcin treatment. Only in pyruvate-grown cells was amino acid uptake strongly inhibited, and the membrane potential was abolished after a short lag time. Also, irreversible killing was more distinct in pyruvate-grown cells. The proton gradient across the cell membrane was only slightly disturbed in both types of cells. Specific inhibitors of the energy metabolism revealed that the high energy state of the membrane was largely supported by hydrolysis of adenosine 5'-triphosphate in glucose-grown cells, whereas the oxidative input through electron transport appeared to be relatively more important in pyruvate-grown cells. Staphylococcin 1580 affected primarily the oxidative energy metabolism, although electron transport is not inhibited. Below a distinct incubation temperature cells were completely resistant to the action of the bacteriocin. Varying the growth temperature had only a slight effect on the transition temperature, but growth in the presence of Tween 80, which greatly enhanced the proportion of unsaturated fatty acids, decreased the transition temperature.

Amino Acids↗

PubMed1985

Effect of various anions on the stability of the coiled coil of skeletal muscle myosin.

The stability of skeletal myosin rod was studied by following the dependence of both papain digestion kinetics and helix-coil transition temperatures on the concentration of neutral salts. The rate of papain-catalyzed digestion of rod to form subfragment 2 and light meromyosin was strongly dependent on chloride concentration but essentially independent of acetate concentration up to 2.0 M. The rod exhibited a biphasic melting curve in 0.6 M NaCl, 5 mM phosphate, and 0.1 mM ethylenediaminetetraacetic acid (EDTA), pH 7.3, with transitions at 45 and 53 degrees C. In 0.6 M CH3COONa, 5 mM phosphate, and 0.1 mM EDTA, pH 7.3, the transitions occurred at 50 and 58 degrees C, respectively. Transition temperatures were obtained with a novel curve-fitting method. The effect of increasing chloride ion concentration on melting profiles was 2-fold. Below 0.6 M salt, the two transition temperatures, Tm,1 and Tm,2, depended on salt concentration such that increasing NaCl concentration caused a small stabilization of the helix while increasing acetate concentration caused the helix to become markedly more stable. Between 0.6 and 1.0 M, variation of chloride concentration had almost no effect on the thermal stability of the rod while increasing acetate concentration increased its stability considerably. Above 1.0 M NaCl, the melting profiles became broad with a third transition being observed (e.g., at 3.0 M, Tm,3 = 38 degrees C), indicating the existence of a region which has a tendency to be destabilized by chloride. The third transition was not observed at comparable concentrations of acetate. This effect of chloride was not expected on the basis of its position in the Hofmeister series.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1985

Protein and lipid structural transitions in cytochrome c oxidase-dimyristoylphosphatidylcholine reconstitutions.

The thermotropic behavior of the mitochondrial enzyme cytochrome c oxidase (EC 1.9.3.1) reconstituted in dimyristoylphosphatidylcholine (DMPC) vesicles has been studied by using high-sensitivity differential scanning calorimetry and fluorescence spectroscopy. The incorporation of cytochrome c oxidase into the phospholipid bilayer perturbs the thermodynamic parameters associated with the lipid phase transition in a manner analogous to other integral membrane proteins: it reduces the enthalpy change, lowers the transition temperature, and reduces the cooperative behavior of the phospholipid molecules. Analysis of the dependence of the enthalpy change on the protein:lipid molar ratio indicates that cytochrome c oxidase prevents 99 +/- 5 lipid molecules from participating in the main gel-liquid-crystalline transition. These phospholipid molecules presumably remain in the same physical state below and above the transition temperature of the bulk lipid, thus providing a more or less constant microenvironment to the protein molecule. The effect of the phospholipid bilayer matrix on the thermodynamic stability of the cytochrome c oxidase complex was examined by high-sensitivity differential scanning calorimetry. Detergent (Tween 80)-solubilized cytochrome c oxidase undergoes a complex, irreversible thermal denaturation process centered at 56 degrees C and characterized by an enthalpy change of 550 +/- 50 kcal/mol of enzyme complex. Reconstitution of the cytochrome c oxidase complex into DMPC vesicles shifts the transition temperature upward to 63 degrees C, indicating that the phospholipid bilayer moiety stabilizes the native conformation of the enzyme. The lipid bilayer environment contributes approximately 10 kcal/mol to the free energy of stabilization of the enzyme complex. The thermal unfolding of cytochrome c oxidase is not a two-state process.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1995

Quantitation of the phase preferences of the major lipids of the Acholeplasma laidlawii B membrane.

We have quantitated the phase preferences of all of the quantitatively significant lipids of fatty acid-homogeneous Acholeplasma laidlawii membranes by determining the effect of small amounts of each lipid on the lamellar/reversed hexagonal phase transition temperature of a phosphatidylethanolamine matrix of identical fatty acid composition using differential scanning calorimetry. We find that the incorporation of small amounts of these lipids produce effects ranging from a moderate depression to a marked elevation of the lamellar/reversed hexagonal phase transition temperature of the corresponding phosphatidylethanolamine. Thus, although the total membrane lipids from this organism form only lamellar phases under physiological conditions, the individual membrane lipids appear to exhibit a wide range of phase preferences. Phosphatidylglycerol and diglucosyldiacylglycerol seem to have relatively strong and weak preferences for the lamellar liquid-crystalline phase, respectively, while monoglucosyldiacylglycerol, and especially acyl polyprenyl glucoside, strongly prefers the reversed hexagonal phase. Most notable in this regard is the phase preference of glycerylphosphoryldiglucosyldiacylglycerol, which strongly destabilizes the reversed hexagonal phase and which we show in the accompanying paper [Lewis, R. N. A. H., & McElhaney, R. N. (1995) Biochemistry 34, 13818-13824] actually prefers the normal micellar phase in isolation. The presence of normal, lamellar, and reversed phase-preferring lipids in a single membrane has important implications for understanding the physical basis of lipid organization and biosynthetic regulation in this and possibly in other organisms. We also show that the characteristic effect of the individual A. laidlawii membrane lipids on the lamellar/reversed hexagonal phase transition temperature of the phosphatidylethanolamine matrix is not well correlated with their polar headgroup intrinsic volumes.(ABSTRACT TRUNCATED AT 250 WORDS)

Acholeplasma laidlawii↗

PubMed1987

Lipid-dependent differential effects of stereoisomers of anesthetic alcohols.

The cis- and trans-alkenols are equally potent general anesthetics but, respectively, lower and raise the gel-to-liquid crystalline phase transition temperature of saturated phosphatidylcholines (Pringle, M.J. and Miller, K.W. (1978) Biochem. Biophys. Res. Commun. 85, 1191-1198). Here we show that although this differential effect is somewhat reduced when a double bond is introduced into the sn-2 position of phosphatidylcholine, it is abolished when the ethanolamine head group is substituted for the choline head group in dimyristoyl lipids at neutral pH. At high pH, however, dimyristoylphosphatidylethanolamine assumes a negative charge, and its phase transition temperature drops to a value close to that for the corresponding phosphatidylcholine. Under these conditions the differential effect of the alkenol isomers is restored; the cis-alkenol lowers, while the trans-alkenol raises, the phase transition temperature of deprotonated dimyristoylphosphatidylethanolamine. Thus, the differential effects of cis- and trans-alkenols on the gel-to-liquid crystalline phase transition are dependent on the physical chemical characteristics of the polar region of the perturbed lipid species, but only weakly on that of the acyl region.

Alcohols↗

PubMedDate not supplied

Influence of low-temperature nucleation on the crystallization process of poly(L-lactide).

The crystallization kinetics of poly(l-lactide), PLLA, is slow enough to allow a quasi-amorphous polymer to be obtained at low temperature simply by quenching from the melt. The PLLA crystallization process was followed by differential scanning calorimetry and optical microscopy after nucleation isothermal treatments at temperatures just below (53 degrees C) and just above (73 degrees C) the glass transition temperature. The crystallization exotherm shown in the heating thermograms shifts toward lower temperatures as the annealing time at 73 degrees C increases. The same effect is shown to a lesser extent when the sample nucleates at 53 degrees C, showing the ability to nucleate in the glassy state, already shown in other polymers. The shape of the DSC thermograms is modeled by using Avrami's theory and allows an estimation of the number of crystallization germs formed. The results of optical microscopy are converted to thermograms by evaluating the average gray level of the image recorded in transmission mode as a function of temperature and calculating its temperature derivative. The shape of such optical thermograms is quite similar to that of the DSC traces but shows some peculiarities after long nucleation treatments. Atomic force microscopy was used to analyze the crystal morphology and is an additional proof of the effect of nucleation in the glassy state. The crystalline morphology observed in samples crystallized after nucleation in the glassy state is qualitatively different from that of samples nucleated above the glass transition temperature, and the number of crystals seems to be much greater than what could be expected from the crystallization kinetics.

Biocompatible Materials↗

PubMed2005

Different effects of long- and short-chain ceramides on the gel-fluid and lamellar-hexagonal transitions of phospholipids: a calorimetric, NMR, and x-ray diffraction study.

The effects on dielaidoylphosphatidylethanolamine (DEPE) bilayers of ceramides containing different N-acyl chains have been studied by differential scanning calorimetry small angle x-ray diffraction and (31)P-NMR spectroscopy. N-palmitoyl (Cer16), N-hexanoyl (Cer6), and N-acetyl (Cer2) sphingosines have been used. Both the gel-fluid and the lamellar-inverted hexagonal transitions of DEPE have been examined in the presence of the various ceramides in the 0-25 mol % concentration range. Pure hydrated ceramides exhibit cooperative endothermic order-disorder transitions at 93 degrees C (Cer16), 60 degrees C (Cer6), and 54 degrees C (Cer2). In DEPE bilayers, Cer16 does not mix with the phospholipid in the gel phase, giving rise to high-melting ceramide-rich domains. Cer16 favors the lamellar-hexagonal transition of DEPE, decreasing the transition temperature. Cer2, on the other hand, is soluble in the gel phase of DEPE, decreasing the gel-fluid and increasing the lamellar-hexagonal transition temperatures, thus effectively stabilizing the lamellar fluid phase. In addition, Cer2 was peculiar in that no equilibrium could be reached for the Cer2-DEPE mixture above 60 degrees C, the lamellar-hexagonal transition shifting with time to temperatures beyond the instrumental range. The properties of Cer6 are intermediate between those of the other two, this ceramide decreasing both the gel-fluid and lamellar-hexagonal transition temperatures. Temperature-composition diagrams have been constructed for the mixtures of DEPE with each of the three ceramides. The different behavior of the long- and short-chain ceramides can be rationalized in terms of their different molecular geometries, Cer16 favoring negative curvature in the monolayers, thus inverted phases, and the opposite being true of the micelle-forming Cer2. These differences may be at the origin of the different physiological effects that are sometimes observed for the long- and short-chain ceramides.

Calorimetry↗

PubMed2005

Acrylamide formation from asparagine under low moisture Maillard reaction conditions. 2. Crystalline vs amorphous model systems.

The formation of acrylamide was investigated in model systems based on asparagine and glucose under low moisture Maillard reaction conditions as a function of reaction temperature, time, physical state, water activity, and glass transition temperature. Equimolar amorphous glucose/asparagine systems with different water activities were prepared by freeze drying and were shown to quickly move to the rubbery state already at room temperature and a water activity of above 0.15. The acrylamide amounts were correlated with physical changes occurring during the reaction. Pyrolysis and kinetics of acrylamide release in amorphous and crystalline glucose/asparagine models indicated the importance of the physical state in acrylamide formation. In amorphous systems, acrylamide was generated in higher concentrations and at lower temperatures as compared to the crystalline samples. Time and temperature are covariant parameters in both systems affecting the acrylamide formation by thermal processes. On the other side, the water activity and glass transition temperature do not seem to be critical parameters for acrylamide formation in the systems studied.

Acrylamide↗

PubMed1979

Properties of bilayer membranes in the phase transition or phase separation region.

The increase in passive permeability of bilayer membranes near the phase transition temperature is usually explained as caused by either the increase in the amount of 'boundary lipid' present in the membrane, or by the increase in lateral compressibility of the membrane. Since both the amount of 'boundary lipid' and the lateral compressibility show a similar anomaly near the transition temperature, it is difficult to distinguish experimentally between the two proposed mechanisms. We have examined some details of both of the proposed pictures. The fluid-solid boundary energy, neglected in previous work, has been computed as a function of the domain size. For a single component uncharged lipid bilayer, the results rule out the existence of even loosely defined solid domains in a fluid phase, or vice versa. Thermodynamic fluctuations, which are responsible for anomalous behaviour near the phase transition temperature, are not intense enough to approximate the formation of a domain of the opposite phase. Turning next to lateral compressibility of bilayer membranes we have considered two-component mixtures in the phase separation region. We present the first calculation of lateral compressibility for such systems. The behaviour shows interesting anomalies, which should correlate with existing and future data on transport across membranes.

Lipid Bilayers↗

PubMed1997

Conformation and thermal denaturation of apocalmodulin: role of electrostatic mutations.

Scanning microcalorimetry and circular dichroism were used to study conformational state and heat denaturation of Ca2+-free synthetic calmodulin (SynCaM) and three charge reversal mutants. We produced evidence for the major role of the electrostatic potential in the stability and flexibility of SynCaM. The substitution of 118DEE120 by 118KKK120 (SynCaM12A) does not influence the flexibility of the protein; the replacement of 82EEE84 by 82KKK84 (SynCaM8) decreases its level, while the combination of these two mutations in SynCaM18A significantly increases the flexibility. The heat denaturation of apoSynCaM and its mutants is well approximated by two two-state transitions with the lower-temperature transition corresponding to C-terminal lobe melting and the higher-temperature one to N-terminal lobe melting. The difference in transition temperatures for the two lobes decreases in SynCaM8 and increases in SynCaM18A, suggesting a modification in the influence of one lobe to the other. The electrostatic mutations change the parameters of thermal denaturation of SynCaM lobes in a similar way as pH conditions affect thermal transition parameters of multidomain proteins, leading to a linear temperature dependence of transition enthalpy. One domain of the N-terminal lobe in apoSynCaM18A is unfolded in the native state. Near-UV CD spectra point out the invariability of the local structure of aromatic residues upon mutations, although the secondary structure undergoes striking transformations. Cacodylate ions strongly and specifically alter the helical content of SynCaM. Our data unambiguously demonstrate that the two lobes are not independent, and interactions between the lobes are mediated by the electrostatic potential of the molecule.

Calcium↗

PubMed1981

Correlation between membrane expansion and temperature-induced membrane fusion.

For each phospholipid membrane, there is a characteristic phase transition temperature, and for each phospholipid spherical membrane, there is a specific 'fusion' temperature. In order to examine the possible correlation between temperature-induced membrane fusion and membrane expansion, the relationship between the physical states of phospholipid membranes at both temperatures have been investigated by the use of the monolayer system. Monolayer expansion studies have indicated that the increase in area per lipid molecule, caused by increasing the temperature from the phase transition to the fusion temperature, is approximately the same for five different phospholipids used. With the same temperature increase, phospholipid monolayers containing cholesterol did not expand appreciably. This correlates qualitatively with the greater inhibition of membrane fusion seen in the spherical phospholipid membrane systems when cholesterol was incorporated in the membrane. The effect of pH on the expansion of phosphatidylserine monolayers was also studied in relation to membrane fusion phenomena. The shift in fusion temperature of the spherical phospholipid membranes due to the change of pH is explained by the shift in phase transition temperatures of lipid membranes. The expanded area per molecule in the monolayer caused by increasing the temperature from the phase transition to the fusion temperature was approximately the same irrespective of surface charge densities.

Cholesterol↗