Pore Model in the Melting Regime of a Lyotropic Biomembrane with an Anionic Phospholipid

Aqueous dispersions of the anionic phospholipid dimyristoyl phosphatidyl glycerol (DMPG) exhibit an unusual “melting regime”, at the phase transition between the ordered (gel) and the disordered (fluid liquid crystal) state of hydrocarbon chains, depending on the ionic strength and DMPG concentration, previously attributed to the pore formation. Dispersions with 150 mM DMPG present a lamellar phase above 23 °C, within the melting regime. In this study, we present a detailed pore model for the analysis of small-angle X-ray scattering (SAXS) results and their variation with temperature, focused on the surface fractions of pores in the bilayers. Large and small toroidal pores are necessary to explain the SAXS results. Pores have DMPG in the fluid conformation, whereas the flat region of the bilayer has DMPG molecules in fluid and in gel conformations. A particular strategy was developed to estimate the charges due to the localization of mobile ions in the system, which is based on the calculation of electron densities by duly considering all molecular and ionic species that characterize the system, and the temperature dependency of their volumes. The best fit to the model of SAXS curves defines that the gel phase transforms initially, at 19.4 °C, in uncoupled bilayers with large pores (radius 93.2 ± 0.5 Å, with water channel diameter 137 ± 1 Å), which transform into small pores along the lamellar phase. The minimum intensity of the SAXS bilayer peak at 30 °C corresponds to a maximum number of small pores, and above 35 °C, the system enters into the normal lamellar fluid phase, without pores. The charge is estimated and shows that the regions with pores contains less Na⁺ ions per polar head; hence, when they are forming, there is a release of Na⁺ ions toward the bulk

Chemical potential differences between the monomer in the

<p><b> state and the monomer in the dissociated “compact” state (</b><b>) calculated from the fractions </b><b> for Conditions A to F as defined in </b><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0049644#pone-0049644-t001" target="_blank"><b>Table 1</b></a><b>.</b> Filled black circles: decamer (); open red circles: octamer (); open green circles: hexamer (); open blue circles: tetramer (); open magenta circles: dimer (); open cyan circles: dissociated “loose” monomer ().</p

Molecular views of <i>Octopus vulgaris</i> hemocyanin in the different aggregation states exploited by QUAFIT to analyse all SAS curves: the “compact” monomer (1) (see Fig. S12 of Ref. [<b>8</b>]) and its hierarchical association products, the dimer (2), the tetramer (4), the hexamer (6), the octamer (8) and the associated full

<p><b>-dimer (10) (see Fig. 3 of Ref. </b><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0049644#pone.0049644-Spinozzi1" target="_blank">[<b>8</b>]</a><b>); the dissociated “loose” monomer (</b><b>) represented by six conformations (see Fig. S11 of Ref. </b><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0049644#pone.0049644-Spinozzi1" target="_blank">[<b>8</b>]</a><b>).</b> The seven rigid domains (a–g) are colour-coded according to Ref. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0049644#pone.0049644-Gatsogiannis1" target="_blank">[15]</a> and the six flexible linkers are shown in dark grey.</p

Batch of SAXS curves with the highest aggregation numbers at pH 7.0.

<p>Each experimental condition is summarised on the right-hand side of the respective SAXS pattern. The entries in column “Experiment” identify each experimental run in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0049644#pone.0049644.s003" target="_blank">Tables S1</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0049644#pone.0049644.s004" target="_blank">S2</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0049644#pone.0049644.s005" target="_blank">S3</a>. The solid black lines are best fitting the experimental data (grey points) normalized to the nominal concentration of the monomer and by the square of its excess scattering length. Notice that the values on the -axis at indicate aggregation numbers. Guinier approximations in the adequate -range are shown as thick blue lines, their extrapolations up to are shown as thick red lines. Histograms on the right-hand side report the protein mass fractions distributed in each aggregation state, as a results of the QUAFIT analysis. Uncertainties on values affect the last decimal place. A zero value is assigned for .</p

SAS curves recorded from <i>Octopus vulgaris</i> hemocyanin samples in four experimental sessions: synchrotron radiation X-rays at LURE (red points

<p><b>–</b><b>) and at the ESRF in two separate sessions (ESRF/1, orange points </b><b>–</b><b> and ESRF/2, cyan points </b><b>–</b><b>); neutrons at the ILL (green points </b><b>–</b><b>).</b> The data are macroscopic differential scattering cross sections, normalized to the nominal concentration of the hemocyanin monomer and to the square of its excess scattering length, upscaled by successive multiples of 10, and reported versus the momentum transfer . Notice that the values on the -axis at represent aggregation numbers. The solid black lines are linear combinations of seven basis functions found by SVD for each of the four sets of curves.</p

SAXS curves referring to <i>Octopus vulgaris</i> hemocyanin in Conditions D, E and F.

<p>Condition D: hemocyanin 5.0 gL in 50 mM glycine pH 9.5, EDTA 5 mM, 100 mM SCN upon F concentration increasing as indicated. Condition E: hemocyanin in 50 mM phosphate pH 7.0, 5 mM sodium dithionite upon protein concentration increasing as indicated. Condition F: hemocyanin in 50 mM Tris/HCl pH 7.0, 5 mM sodium dithionite upon protein concentration increasing as indicated. Results of the Guinier analysis for these conditions are reported in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0049644#pone-0049644-t001" target="_blank">Table 1</a>. See the caption of Fig. 1 for more details.</p

Small-Angle Neutron Scattering Reveals the Nanostructure of Liposomes with Embedded OprF Porins of Pseudomonas aeruginosa

The use of liposomes as drug delivery systems emerged in the last decades in view of their capacity and versatility to deliver a variety of therapeutic agents. By means of small-angle neutron scattering (SANS), we performed a detailed characterization of liposomes containing outer membrane protein F (OprF), the main porin of the Pseudomonas aeruginosa bacterium outer membrane. These OprF-liposomes are the basis of a novel vaccine against this antibiotic-resistant bacterium, which is one of the main hospital-acquired pathogens and causes each year a significant number of deaths. SANS data were analyzed by a specific model we created to quantify the crucial information about the structure of the liposome containing OprF, including the lipid bilayer structure, the amount of protein in the lipid bilayer, the average protein localization, and the effect of the protein incorporation on the lipid bilayer. Quantification of such structural information is important to enhance the design of liposomal delivery systems for therapeutic applications

Structural and Thermodynamic Properties of Nanoparticle–Protein Complexes: A Combined SAXS and SANS Study

We propose a novel method for determining the structural and thermodynamic properties of nanoparticle–protein complexes under physiological conditions. The method consists of collecting a full set of small-angle X-ray and neutron-scattering measurements in solutions with different concentrations of nanoparticles and protein. The nanoparticle–protein dissociation process is described in the framework of the Hill cooperative model, based on which the whole set of X-ray and neutron-scattering data is fitted simultaneously. This method is applied to water solutions of gold nanoparticles in the presence of human serum albumin without any previous manipulation and can be, in principle, extended to all systems. We demonstrate that the protein dissociation constant, the Hill coefficient, and the stoichiometry of the nanoparticle–protein complex are obtained with a high degree of confidence