Thermotropic and barotropic phase transitions of N-methylated dipalmitoylphosphatidylethanolamine bilayers

In order to understand the effect of polar head group modification on the thermotropic and barotropic phase behavior of phospholipid bilayer membranes, the phase transitions of dipalmitoylphosphatidylethanolamine (DPPE), dipalmitoylphosphatidyl-N-methylethanolamine (DPMePE), dipalmitoylphosphatidyl-N,N-dimethylethanolamine (DPMe2PE) and dipalmitoylphosphatidylcholine (DPPC) bilayer membranes were observed by differential scanning calorimetry and high-pressure optical methods. The temperatures of the so-called main transition from the gel (L(beta)) or ripple gel (P(beta)') phase to the liquid crystalline (L(alpha)) phase were almost linearly elevated by applying pressure. The slope of the temperature-pressure boundary, dT/dp, was in the range of 0.220-0.264 K MPa(-1) depending on the number of methyl groups in the head group of lipids. The main-transition temperatures of N-methylated DPPEs decreased with increasing size of head group by stepwise N-methylation. On the other hand, there was no significant difference in thermodynamic quantities of the main transition between the phospholipids. With respect to the transition from the subgel (L(c)) phase to the lamellar gel (L(beta) or L(beta)') phase, the transition temperatures were also elevated by applying pressure. In the case of DPPE bilayer the L(c)/L(beta) transition appeared at a pressure higher than 21.8 MPa. At a pressure below 21.8 MPa the L(c)/L(alpha) transition was observed at a temperature higher than the main-transition temperature. The main (L(beta)/L(alpha)) transition can be recognized as the transformation between metastable phases in the range from ambient pressure to 21.8 MPa. Polymorphism in the gel phase is characteristic of DPPC bilayer membrane unlike other lipid bilayers used in this study: the L(beta)', P(beta)' and pressure-induced interdigitated gel (L(beta)I) phases were observed only in the DPPC bilayer. Regarding the bilayers of DPPE, DPMePE and DPMe2PE, the interdigitation of acyl chain did not appear even at pressures as high as 200 MPa.     

Synthesis and characterization of betaine-like diacyl lipids: zwitterionic lipids with the cationic amine at the bilayer interface

We synthesized and characterized a series of zwitterionic, acetate-terminated, quaternized amine diacyl lipids (AQ). These lipids have an inverted headgroup orientation as compared to naturally occurring phosphatidylcholine (PC) lipids; the cationic group is anchored at the membrane interface, while the anionic group extends into the aqueous phase. AQ lipids preferentially interact with highly polarizable anions (ClO(4)(-)) over less polarizable ions (Cl(-)), in accord with the Hofmeister series, as measured by the change in zeta potential of AQ liposomes. Conversely, AQ lipids have a weaker association with calcium than do PC lipids. The transition temperatures (Tm) of the AQ lipids are similar to the Tm observed with phosphatidylethanolamine (PE) lipids of the same chain length. AQ lipids form large lipid sheets after heating and sonication; however, in the presence of cholesterol (Chol), these lipids form stable liposomes that encapsulate carboxyfluorescein. The AQ:Chol liposomes retain their contents in the presence of serum at 37°C, and when injected intravenously into mice, their organ biodistribution is similar to that observed with PC:Chol liposomes. AQ lipids demonstrate that modulating the headgroup charge orientation significantly alters the biophysical properties of liposomes. For the drug carrier field, these new materials provide a non-phosphate containing zwitterlipid for the production of lipid vesicles.     

Interaction of the pore-forming protein equinatoxin II with model lipid membranes: A calorimetric and spectroscopic study.

The interactions of equinatoxin II (EqTxII) with zwitterionic (DPPC) and anionic (DPPG) phospholipids and an equimolar mixture of the two phospholipids (DPPC/DPPG) have been investigated by differential scanning calorimetry (DSC), CD-spectropolarimetry, intrinsic emission fluorescence spectroscopy, and ultrasonic velocimetry. EqTxII binds to small unilamellar vesicles formed from negatively charged DPPG lipids, causing a marked reduction in the cooperativity and enthalpy of their gel/liquid-crystalline phase transition. This transition is completely abolished at a lipid-to-protein ratio, L/P, of 10. For the mixed DPPC/DPPG vesicles, a 2-fold greater lipid-to-protein ratio (L/P = 20) is required to abolish the phase transition, which corresponds to the same negative charge (-10) of lipid molecules per EqTxII molecule. The disappearance of the phase transition of the lipids apparently corresponds to the precipitation of the lipid-protein complex, as suggested by our sound velocity measurements. Based on the far-UV CD spectra, EqTxII undergoes two structural transitions in the presence of negatively charged vesicles (DPPG). The first transition coincides with the gel/liquid-crystalline phase transition of the lipids, which suggests that the liquid-crystalline form of negatively charged lipids triggers structural changes in EqTxII. The second transition involves the formation of alpha-helical structure. Based on these observations, we propose that, in addition to electrostatic interactions, hydrophobic interactions play an important role in EqTxII-membrane association.     

Sequence-dependent cytotoxicity of second-generation oligonucleotides

In this study, we have examined the potential of second-generation antisense chimeric 2′-O-(2-methoxy)ethyl/DNA phosphorothioate oligonucleotides (ONs) to affect cell growth through non-antisense mechanisms. Evaluation of a series of ONs demonstrated that only a small number were cytotoxic at concentrations close to those required for antisense activity. Toxicity of the ONs appeared to be sequence dependent and could be affected by base and backbone modifications. Caspase-3 activation occurs with some ONs and it is most likely secondary to necrosis rather than apoptosis, since cells treated with toxic ONs did not show chromatin condensation, but did exhibit high-extracellular lactate dehydrogenase activity. Caspase-3 activation does not correlate with and appears not to be required for the inhibition of cell proliferation. Toxicity was only observed when ONs were delivered intracellularly. The mechanism by which one of the most cytotoxic ON produces cytotoxicity was investigated in more detail. Treatment with the cytotoxic ON caused disruption of lysosomes and Pepstatin A, a specific inhibitor of aspartic proteases, reduced the cytotoxicity of the ON. Reduction of lysosomal aspartic protease cathepsin D by prior treatment with cathepsin D-specific antisense ON did not attenuate the cytotoxicity, suggesting that other aspartic proteases play a crucial role in the cellular proliferation inhibition by ONs.     

Interaction between artificial membranes and enflurane,a general volatile anesthetic: DPPC-enflurane interaction

The structural modifications of the dipalmitoylphosphatidylcholine (DPPC) organization induced by increasing concentration of the volatile anesthetic enflurane have been studied by differential scanning calorimetry, small-angle, and wide-angle x-ray scattering. The interaction of enflurane with DPPC depends on at least two factors: the enflurane-to-lipid concentration ratio and the initial organization of the lipids. At 25 degrees C (gel state), the penetration of enflurane within the lipids induces the apparition of two different mixed lipid phases. At low anesthetic-to-lipid molar ratio, the smectic distance increases whereas the direction of the chain tilt changes from a tilt toward next-neighbors to a tilt between next-neighbors creating a new gel phase called L(beta')(2NNN). At high ratio, the smectic distance is much smaller than for the pure L(beta') DPPC phase, i.e., 50 A compared to 65 A, the aliphatic chains are perpendicular to the membrane and the fusion temperature of the phase is 33 degrees C. The electron profile of this phase that has been called L(beta)(i), indicates that the lipids are fully interdigitated. At 45 degrees C (fluid state), a new melted phase, called L(alpha)(2), was found, in which the smectic distance decreased compared to the initial pure L(alpha)(1) DPPC phase. The thermotropic behavior of the mixed phases has also been characterized by simultaneous x-ray scattering and differential scanning calorimetry measurements using the Microcalix calorimeter of our own. Finally, titration curves of enflurane effect in the mixed lipidic phase has been obtained by using the fluorescent lipid probe Laurdan. Measurements as a function of temperature or at constant temperature, i.e., 25 degrees C and 45 degrees C give, for the maximal effect, an enflurane-to-lipid ratio (M/M), within the membrane, of 1 and 2 for the L(alpha)(2) and the L(beta)(i) lamellar phase respectively. All the results taken together allowed to draw a pseudo-binary phase diagram of enflurane-dipalmitoylphosphatidylcholine in excess water.     

Phospholipid-cationic lipid interactions: influences on membrane and vesicle properties

Liposomes composed of synthetic dialkyl cationic lipids and zwitterionic phospholipids such as dioleoylphosphatidylethanolamine have been studied extensively as vehicles for gene delivery, but the broader potentials of these cationic liposomes for drug delivery have not. An understanding of phospholipid-cationic lipid interactions is essential for rational development of this potential. We evaluated the effect of the cationic lipid DOTAP (N-[1-(2,3-dioleoyloxy)propyl]-N,N,N-trimethylammonium) on liposome physical properties such as size and membrane domain structure. DSC (differential scanning calorimetry) showed progressive decrease and broadening of the phase transition temperature of dipalmitoylphosphatidylcholine (DPPC) with increasing fraction of DOTAP, in the range of 0.4-20 mol%. Laurdan (6-dodecanolyldimethylamino-naphthalene), a fluorescent probe of membrane domain structure, showed that DOTAP and DPPC remained miscible at all ratios tested. DOTAP reduced the size of spontaneously-forming PC-containing liposomes, regardless of the acyl chain length and degree of saturation. The anionic lipid DOPG (dioleoylphosphatidylglycerol) had similar effects on DPPC membrane fluidity and size. However, DOTAP/DOPC (50/50) vesicles were taken up avidly by OVCAR-3 human ovarian tumor cells, in contrast to DOPG/DOPC (50/50) liposomes. Overall, DOTAP exerts potent effects on bilayer physical properties, and may provide advantages for drug delivery.     

Effect of hydrophobic structure on the catalysis of nitric oxide release from zwitterionic diazeniumdiolates in surfactant and liposome media.

The effect of phospholipid liposomes and surfactant micelles on the rate of nitric oxide release from zwitterionic diazeniumdiolates, R1R2N[N(O)NO]-, with significant hydrophobic structure, has been explored. The acid-catalyzed dissociation of NO has been examined in phosphate-buffered solutions of sodium dodecylsulfate (SDS) micelles and 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and 1,2-dipalmitoyl-sn-glycero-3-[phospho-(1-glycerol)] sodium salt (DPPG) phospholipid liposomes. The reaction behavior of dibenzylamine-, monobenzylamine-, and dibutylamine-derived substrates [1]: R1 = C6H5CH2, R2 = C6H5CH2 NH2+(CH2)2, 2: R1 = C6H5CH2, R2 = NH3+(CH2)2, and 3: R1 = n-butyl, R2 = n-butyl-NH2+(CH2)6] has been compared with that of SPER/NO, 4: R1 = H2N(CH2)3, R2 = H2N(CH2) 3NH2+(CH2)4]. Catalysis of NO release is observed in both micellar and liposome media. Hydrophobic interactions contribute to micellar binding for 1-3 and appear to be the main factor facilitating catalysis by charge neutral DPPC liposomes. Binding constants for the association of 1 and 3 with SDS micelles were 3-fold larger than those previously obtained with comparable zwitterionic substrates lacking their hydrophobic structure. Anionic DPPG liposomes were much more effective in catalyzing NO release than either DPPC liposomes or SDS micelles. DPPG liposomes (at 10 mM total lipid) induced a 30-fold increase in the NO dissociation rate of SPER/NO compared to 12- and 14-fold increases in that of 1 and 3.     

Interplay of polyethyleneimine molecular weight and oligonucleotide backbone chemistry in the dynamics of antisense activity

The widespread utilization of gene silencing techniques, such as antisense, is impeded by the poor cellular delivery of oligonucleotides (ONs). Rational design of carriers for enhanced ON delivery demands a better understanding of the role of the vector on the extent and time course of antisense effects. The aim of this study is to understand the effects of polymer molecular weight (MW) and ON backbone chemistry on antisense activity. Complexes were prepared between branched polyethyleneimine (PEI) of various MWs and ONs of phosphodiester and phosphorothioate chemistries. We measured their physico-chemical properties and evaluated their ability to deliver ONs to cells, leading to an antisense response. Our key finding is that the antisense activity is not determined solely by PEI MW or by ON chemistry, but rather by the interplay of both factors. While the extent of target mRNA down-regulation was determined primarily by the polymer MW, dynamics were determined principally by the ON chemistry. Of particular importance is the strength of interactions between the carrier and the ON, which determines the rate at which the ONs are delivered intracellularly. We also present a mathematical model of the antisense process to highlight the importance of ON delivery to antisense down-regulation.     

Effect of progesterone on DPPC membrane: evidence for lateral phase separation and inverse action in lipid dynamics

Interactions of progesterone with zwitterionic dipalmitoyl phosphatidylcholine (DPPC) multilamellar liposomes were investigated as a function of temperature and progesterone concentration by using three non-invasive techniques namely Fourier transform infrared spectroscopy, turbidity at 440 nm, and differential scanning calorimetry. The results reveal that progesterone changes the physical properties of DPPC bilayers by decreasing the main phase-transition temperature, abolishing the pre-transition, broadening the phase-transition profile, disordering the system both in gel and liquid crystalline phase, increasing the dynamics at low concentrations whereas stabilizing the membrane at high concentrations, and inducing phase separation. Progesterone does not change the hydration of the CO groups, while it strengthens the hydrogen bonding between the PO2- groups of lipids and the water molecules around.     

Titration calorimetric and differential scanning calorimetric studies of the interactions of n-butanol with several phases of dipalmitoylphosphatidylcholine.

The interactions of n-butanol with dipalmitoylphosphatidylcholine (DPPC) were studied using titration calorimetry and differential scanning calorimetry (DSC). DSC results indicated that n-butanol induces the interdigitated phase in DPPC above 10 mg/mL butanol. A new application of titration calorimetry for measuring partition coefficients of nonsaturating solutes into lipids was developed. The partition coefficients and the heat of binding of n-butanol into DPPC were measured for the L beta', P beta', L alpha, and L beta I phases of DPPC. The partition coefficients were temperature dependent and ranged from 70 to 110 for the L beta I phase, from 170 to 183 for the L alpha phase, and similar to that for the L beta I phase in the P beta' phase. The binding to the L beta' phase could not be detected, giving an upper limit for this partition coefficient of 23. The enthalpies for binding to the L beta I and L alpha phases were 1.0 and 1.5 kcal/mol, respectively. The van't Hoff enthalpy was in good agreement with the calorimetric enthalpy for the partitioning into the L alpha phase; however, it was greater than the calorimetric enthalpy for the L beta I phase, suggesting that the interaction of n-butanol with this phase is cooperative in some way.     

Association of acylated cationic decapeptides with dipalmitoylphosphatidylserine-dipalmitoylphosphatidylcholine lipid membranes.

The interaction of three acylated and cationic decapeptides with lipid membranes composed of dipalmitoylphosphatidylcholine (DPPC) and dipalmitoylphosphatidylserine (DPPS) has been studied by means of fluorescence spectroscopy and differential scanning calorimetry (DSC). The synthetic model decapeptides that are N-terminally linked with C(2), C(8), and C(14) acyl chains contain four basic histidine residues in their identical amino acid sequence. A binding model, based on changes in the intrinsic fluorescent properties of the peptides upon association with the DPPC-DPPS membranes, is used to estimate the peptide-membrane dissociation constants. The results clearly show that all three peptides have a higher affinity to liposomes containing DPPS lipids due to non-specific electrostatic interactions between the cationic peptides and the anionic DPPS lipids. Furthermore, it is found that the acyl chain length of the peptides plays a crucial role for the binding. A preference for fluid phase membranes as compared to gel phase membranes is generally observed for all three peptides. DSC is used to characterise the influence of the three peptides on the thermodynamic phase behaviour of the binary DPPC-DPPS lipid mixture. The extent of peptide association deduced from the heat capacity measurements suggests a strong binding and membrane insertion of the C(14) acylated peptide in accordance with the fluorescence measurements.     

The phase transition behavior of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) model membrane influenced by 2,4-dichlorophenol--an FT-Raman spectroscopy study

The effect of 2,4-dichlorophenol (DCP) on the structures and phase transitions of fully hydrated 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) liposomes was studied using FT-Raman spectroscopy. Whereas the Raman frequency shifts of the most frequently investigated bands of C-C and C-H stretching regions only indicate the main phase transition (P(beta')-L(alpha)) of the pure DPPC/water system, the Raman shift of C-H scissoring vibration at 1440 cm(-1) was found to be able to reveal the pretransition (L(beta')-P(beta')) as well. Analyzing the spectral parameters of the trans band at 1128 cm(-1), which does not overlap with DCP vibrational modes, a continuous decrease of trans conformations was found with increasing DCP concentration at 26 degrees C accompanying the phase transitions L(beta')-P(beta') and P(beta')-L(alpha). The intensity ratio of the symmetrical and asymmetrical methylene stretching bands (at 2850 cm(-1) and 2880 cm(-1)), defined as the disorder parameter by Levin [Levin, I.W., 1985. Two types of hydrocarbon chain interdigitation in sphingomielin bilayers. Biochemistry 24, 6282-6286], indicated that in the interdigitated phase (L(I)) the order is markedly high and comparable with that of L(beta). Both the phase transition P(beta')-L(alpha) in the DCP/DPPC molar ratio range of 10/100-50/100 and the phase transition L(I)-L(alpha) led to a significant increase of disordered chains and the presence of DCP molecules induced a more disordered chain region than that observed in the L(alpha) phase of DPPC. Nevertheless, it was found that the L(alpha) phase with DCP contains approximately the same amount of trans conformers than that without DCP.     

Interactions of ciprofloxacin with DPPC and DPPG: fluorescence anisotropy, ATR-FTIR and 31P NMR spectroscopies and conformational analysis

The interactions between a drug and lipids may be critical for the pharmacological activity. We previously showed that the ability of a fluoroquinolone antibiotic, ciprofloxacin, to induce disorder and modify the orientation of the acyl chains is related to its propensity to be expelled from a monolayer upon compression [1]. Here, we compared the binding of ciprofloxacin on DPPC and DPPG liposomes (or mixtures of phospholipids [DOPC:DPPC], and [DOPC:DPPG]) using quasi-elastic light scattering and steady-state fluorescence anisotropy. We also investigated ciprofloxacin effects on the transition temperature (T(m)) of lipids and on the mobility of phosphate head groups using Attenuated Total Reflection Fourier Transform Infrared-Red Spectroscopy (ATR-FTIR) and (31)P Nuclear Magnetic Resonance (NMR) respectively. In the presence of ciprofloxacin we observed a dose-dependent increase of the size of the DPPG liposomes whereas no effect was evidenced for DPPC liposomes. The binding constants K(app) were in the order of 10(5) M(-1) and the affinity appeared dependent on the negative charge of liposomes: DPPG>DOPC:DPPG (1:1; M:M)>DPPC>DOPC:DPPC (1:1; M:M). As compared to the control samples, the chemical shift anisotropy (Deltasigma) values determined by (31)P NMR showed an increase of 5 and 9 ppm for DPPC:CIP (1:1; M:M) and DPPG:CIP (1:1; M:M) respectively. ATR-FTIR experiments showed that ciprofloxacin had no effect on the T(m) of DPPC but increased the order of the acyl chains both below and above this temperature. In contrast, with DPPG, ciprofloxacin induced a marked broadening effect on the transition with a decrease of the acyl chain order below its T(m) and an increase above this temperature. Altogether with the results from the conformational analysis, these data demonstrated that the interactions of ciprofloxacin with lipids depend markedly on the nature of their phosphate head groups and that ciprofloxacin interacts preferentially with anionic lipid compounds, like phosphatidylglycerol, present at a high content in these membranes.     

Lipid transfer between cationic vesicles and lipid-DNA lipoplexes: Effect of serum

Differential scanning calorimetry was used to examine the lipid exchange between model lipid systems, including vesicles of the cationic lipoids ethyldimyristoylphosphatidylcholine (EDMPC), ethyldipalmitoylphosphatidylcholine (EDPPC) or their complexes with DNA (lipoplexes), and the zwitterionic lipids (DMPC, DPPC). The changes of the lipid phase transition parameters (temperature, enthalpy, and cooperativity) upon consecutive temperature scans was used as an indication of lipid mixing between aggregates. A selective lipid transfer of the shorter-chain cationic lipoid EDMPC into the longer-chain aggregates was inferred. In contrast, transfer was hindered when EDMPC (but not EDPPC) was bound to DNA in the lipoplexes. These data support a simple molecular lipid exchange mechanism, but not lipid bilayer fusion. Exchange via lipid monomers is considerably more facile for the cationic ethylphosphatidylcholines than for zwitterionic phosphatidylcholines, presumably due to the higher monomer solubility of the charged lipids. With the cationic liposomes, lipid transfer was strongly promoted by the presence of serum in the dispersing medium. Serum proteins are presumed to be responsible for the accelerated transfer, since the effect was strongly reduced upon heating the serum to 80 °C. The effect of serum indicates that even though much lipoplex lipid is inaccessible due to the multilayered structure, the barrier due to buried lipid can be easily overcome. Serum did not noticeably promote the lipid exchange of zwitterionic liposomes. The phenomenon is of potential importance for the application of cationic liposomes to nonviral gene delivery, which often involves the presence of serum in vitro, and necessarily involves serum contact in vivo.     

Interaction of selected anthocyanins with erythrocytes and liposome membranes

Anthocyanins are one of the main flavonoid groups. They are responsible for, e.g., the color of plants and have antioxidant features and a wide spectrum of medical activity. The subject of the study was the following compounds that belong to the anthocyanins and which can be found, e.g., in strawberries and chokeberries: callistephin chloride (pelargonidin-3-O-glucoside chloride) and ideain chloride (cyanidin-3-O-galactoside chloride). The aim of the study was to determine the compounds’ antioxidant activity towards the erythrocyte membrane and changes incurred by the tested anthocyanins in the lipid phase of the erythrocyte membrane, in liposomes composed of erythrocyte lipids and in DPPC, DPPC/cholesterol and egg lecithin liposomes. In particular, we studied the effect of the two selected anthocyanins on red blood cell morphology, on packing order in the lipid hydrophilic phase, on fluidity of the hydrophobic phase, as well as on the temperature of phase transition in DPPC and DPPC/cholesterol liposomes. Fluorimetry with the Laurdan and Prodan probes indicated increased packing density in the hydrophilic phase of the membrane in the presence of anthocyanins. Using the fluorescence probes DPH and TMA-DPH, no effect was noted inside the hydrophobic phase of the membrane, as the lipid bilayer fluidity was not modified. The compounds slightly lowered the phase transition temperature of phosphatidylcholine liposomes. The study has shown that both anthocyanins are incorporated into the outer region of the erythrocyte membrane, affecting its shape and lipid packing order, which is reflected in the increasing number of echinocytes. The investigation proved that the compounds penetrate only the outer part of the external lipid layer of liposomes composed of erythrocyte lipids, DPPC, DPPC/cholesterol and egg lecithin lipids, changing its packing order. Fluorimetry studies with DPH-PA proved that the tested anthocyanins are very effective antioxidants. The antioxidant activity of the compounds was comparable with the activity of Trolox®.     

Hybridization of antisense oligonucleotides with alpha-sarcin loop region of Escherichia coli 23S rRNA

Binding of complementary oligonucleotides (ONs) with alpha-sarcin loop region (2638-2682) of Escherichia coli 23S rRNA was investigated. Four of the tested pentadecanucleotides efficiently bound to target sequences with association rate and equilibrium constants approximately 10(3) M(-1)s(-1) and 10(7) M(-1), respectively. ON S5 (CGAGAGGACCGGAGU) complementary to the sequence 2658-2672 displayed the highest affinity to the target. Activation energy for binding of ON S5 was measured to be 11 kcal/mol; this value corresponds to approximately 10% of the calculated enthalpy of the local RNA structure unfolding in the presence of this oligonucleotide. The activation energy value is evidence for the heteroduplex formation to occur via strand displacement pathway; the initiation of heteroduplex formation requires disruption of 1-2 base pairs in RNA hairpin.     

Mapping of HIV-1 integrase preferences for target site selection with various oligonucleotides

HIV integrase (IN) catalyzes the insertion of proviral DNA into the host cell chromosome. While IN has strict sequence requirements for the viral cDNA ends, the integration site preference has been shown to be very diverse. Here, we mapped the HIV IN strand transfer reaction requirements using various short oligonucleotides (ON) that mimic the target DNA. Most double stranded DNA dodecamers served as excellent IN targets with variable integration efficiency depending mostly on the ON sequences. The preferred integration was lost with any changes in the geometry of the DNA double helical structures. Various hairpin-loop-forming ONs also served as efficient integration targets. Similar integration preferences were also observed for ONs, in which the nucleotide hairpin loop was replaced with a flexible aliphatic linker. The integration biases with all target DNA structures tested were significantly influenced by changes in the resulting secondary ON structures.     

The hydration pressure between lipid bilayers. Comparison of measurements using x-ray diffraction and calorimetry.

The hydration pressure between dipalmitoyl phosphatidyl-N,N-dimethylethanolamine (DPPE-Me2) bilayers has been analyzed by both x-ray diffraction measurements of osmotically stressed liposomes and by differential scanning calorimetry. By the x-ray method, we obtain a magnitude (Po) and decay length (lambda) for the hydration pressure which are both quite similar to those found for bilayers of other zwitterionic lipids, such as phosphatidylcholines. That is, x-ray analysis of DPPE-Me2 in the gel phase gives lambda = 1.3 A, the same as that previously measured for the analogous gel phase lipid dipalmitoylphosphatidylcholine (DPPC), and Po = 3.9 x 10(9) dyn/cm2, which is in excellent agreement with the value of 3.6 x 10(9) dyn/cm2 calculated from the measured Volta potential of DPPE-Me2 monolayers in equilibrium with liposomes. These results indicate that the removal of one methyl group to convert DPPC to DPPE-Me2 does not markedly alter the range or magnitude of the hydration pressure. Calorimetry shows that the main gel to liquid-crystalline phase transition temperature of DPPE-Me2 is approximately constant for water contents ranging from 80 to 10 water molecules per lipid molecule, but increases monotonically with decreasing water content below 10 waters per lipid. A theoretical fit to these temperature vs. water content data predicts lambda = 6.7 A. The difference in observed values of lambda for x-ray and calorimetry measurements can be explained by effects on the thermograms of additional intra- and intermolecular interactions which occur at low water contents where apposing bilayers are in contact.(ABSTRACT TRUNCATED AT 250 WORDS)     

High pressure effect on the main transition from the ripple gel P'beta phase to the liquid crystal (Lalpha) phase in dipalmitoylphosphatidylcholine. Microcalorimetric study

Scanning microcalorimetry has been used to study the high pressure effect on the main transition from the ripple gel P'(beta) phase to the liquid crystal (L(alpha)) phase in DPPC (dipalmitoylphosphatidylcholine). It has been demonstrated that an increase of the pressure by 200 MPa shifts the transition to higher temperatures by 36.4 degrees. The pressure increase does not affect the cooperativity of transition but reduces noticeably its enthalpy. The changes of the molar partial volume, isothermal compressibility as well as volume thermal expansibility during transition in DPPC suspension have been estimated. It has been shown that monovalent ions (Na(+), Cl(-)) in solution slightly affect the main thermodynamic parameters of the transition. Calcium ions significantly decrease distinction in compressibility and thermal expansibility between liquid-crystal and ripple gel phases of lipid suspension, which in its turn reflects less difference in their volume fluctuations.