Biomedical subjects

P Laggner

Publications and source records attributed to P Laggner.

At least 19 recordsLinked to original sources

PubMed2005

In situ tensile testing of human aortas by time-resolved small-angle X-ray scattering.

The collagen diffraction patterns of human aortas under uniaxial tensile test conditions have been investigated by synchrotron small-angle X-ray scattering. Using a recently designed tensile testing device the orientation and d-spacing of the collagen fibers in the adventitial layer have been measured in situ with the macroscopic force and sample stretching under physiological conditions. The results show a direct relation between the orientation and extension of the collagen fibers on the nanoscopic level and the macroscopic stress and strain. This is attributed first to a straightening, second to a reorientation of the collagen fibers, and third to an uptake of the increasing loads by the collagen fibers.

Aorta↗

PubMed2005

Effects of N-alkyl-N,N-dimethylamine-N-oxides on the activity of purified sarcoplasmic reticulum Ca(2+)-transporting ATPase.

N-alkyl-N,N-dimethylamine-N-oxides (CnNO, n = 10-20 is the number of alkyl carbon atoms) stimulate the skeletal sarcoplasmic reticulum (SR) Ca(2+)-transporting ATPase activity at low concentrations and inhibit it at high concentrations. The minimum concentration (cmin), at which CnNO inhibits the ATPase, continuously decreases up to n = 16-18 and then increases. The values of Cmin are smaller than the CnNO critical micelle concentration (cmc) for C10NO-C14NO homologs, but larger than cmc for C18NO-C20NO homologs. The ATPase inhibition is caused by the CnNO-induced lipid bilayer structural perturbation in the ATPase annular region, modulated by the partition equilibria of the CnNO molecules between the bilayer and aqueous phase for short alkyl chain (n = 10-16) CnNO homologs, and between the bilayer, micelles and aqueous phase for long alkyl chain (n = 18-20) CnNO homologs.

Animals↗

PubMed2004

Structure and fluctuations of phosphatidylcholines in the vicinity of the main phase transition.

We have determined the structural properties and bending fluctuations of fully hydrated phosphatidylcholine multibilayers in the fluid (Lalpha) phase, as well as the structure of the ripple (Pbeta') phase near the main phase transition temperature (TM) by x-ray diffraction. The number of carbons, nHC, per acyl chain of the studied disaturated lipids varied from 14 to 22. All lipids exhibit a nonlinear increase of the lamellar repeat distance d in the Lalpha phase upon approaching TM, known as "anomalous swelling." The nonlinear increase reduces with chain length, but levels off at a constant value of about 0.5 A for lipids with more than 18 hydrocarbons per chain. A detailed analysis shows that anomalous swelling has two components. One is due to an expansion of the water layer, which decreases with chain length and finally vanishes for nHC >18. The second component is due to a bilayer thickness increase, which remains unchanged in its temperature dependence, including a nonlinear component of about 0.5 A in the vicinity of TM. Thus, anomalous swelling above 18 hydrocarbons per chain is due to the pretransitional effects on the membrane only. These results are supported by a bending fluctuation analysis revealing increased undulations close to TM only for the short chain lipids. We have further calculated the electron density maps in the ripple phase and find no coupling of the magnitude of the ripple amplitude to the chain length effects observed in the Lalpha phase. Hence, in agreement with an earlier report by Mason et al. [Phys. Rev. E 63, 030902 (2001)] there is no connection between the formation of the ripple phase and anomalous swelling.

Biophysics↗

PubMed2004

Efforts to understand the molecular basis of hypertension through drug:membrane interactions.

Biological membranes play an essential role in the drug action. They constitute the first barrier for drugs to exert their biological action. AT1 antagonists are amphiphilic molecules and are hypothesized to act on AT1 receptor through incorporation (first step) and lateral diffusion through membrane bilayers (second step). Various biophysical methods along with Molecular Modelling were applied in order to explore the plausible two step proposed mechanism of action for this class of antihypertensive drugs.

Amino Acids↗

PubMed2001

Mechanism of the interaction of beta(2)-glycoprotein I with negatively charged phospholipid membranes.

In an attempt to understand the multifunctional involvement of beta(2)-glycoprotein I (beta(2)GPI) in autoimmune diseases, thrombosis, atherosclerosis, and inflammatory processes, substantial interest is focused on the interaction of beta(2)GPI with negatively charged ligands, in particular, with acidic phospholipids. In this study, unilamellar vesicles composed of cardiolipin were used as in vitro membrane system to test and further refine a model of interaction based on the crystal structure of beta(2)GPI. The data suggest that beta(2)GPI anchors to the membrane surface with its hydrophobic loop adjacent to the positively charged lysine rich region in domain V. Subsequently, beta(2)GPI penetrates the membrane interfacial headgroup region as indicated by a restriction of the lipid side chain mobility, but without formation of a nonbilayer lipid phase. A structural rearrangement of beta(2)GPI upon lipid binding was detected by microcalorimetry and may result in the exposure of cryptic epitopes located in the complement control protein domains. This lipid-dependent conformational change may induce oligomerization of beta(2)GPI and promote intermolecular associations. Thus, the aggregation tendency of beta(2)GPI may serve as the basis for the formation of a molecular link between cells but may also be an essential feature for binding of autoantibodies and hence determine the role of beta(2)GPI in autoimmune diseases.

Apolipoproteins↗

PubMed2001

Differential membrane fluidization by active and inactive cannabinoid analogues.

The effects of the two cannabinomimetic drugs (-)-2-(6a,7,10,10a-tetrahydro-6,6,9-trimethyl-1-hydroxy-6H-dibenzo[b,d]pyranyl-2-(hexyl)-1,3-dithiolane (AMG-3) and its pharmacologically less active 1-methoxy analogue (AMG-18) on the thermotropic and structural properties of dipalmitoyl-sn-glycero-3-phosphorylcholine (DPPC) liposomes have been studied by X-ray diffraction and differential scanning calorimetry (DSC). DSC data revealed that the incorporation of the drugs affect differently the thermotropic properties of DPPC. The presence of the more active drug distinctly broadened and attenuated both the pretransition and main phase transition of DPPC bilayers, while the inactive analogue had only minor effects. Small and wide angle X-ray diffraction data showed that the two cannabinoids have different effects on the lipid phase structures and on the hydrocarbon chain packing. The pharmacologically active analogue, AMG-3, was found to efficiently fluidize domains of the lipids in the L(beta)' gel phase, and to perturb the regular multibilayer lattice. In the liquid crystalline L(alpha) phase, AMG-3 was also found to cause irregularities in packing, suggesting that the drug induces local curvature. At the same concentration, the inactive AMG-18 had only minor structural effects on the lipids. At about 10-fold or higher concentrations, AMG-18 was found to produce similar but still less pronounced effects in comparison to those observed by AMG-3. The dose-dependent, different thermotropic and structural effects by the two cannabinoid analogues suggest that these may be related to their biological activity.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed2000

Self-assembly of large, ordered lamellae from non-bilayer lipids and integral membrane proteins in vitro.

In many biological membranes, the major lipids are "non-bilayer lipids," which in purified form cannot be arranged in a lamellar structure. The structural and functional roles of these lipids are poorly understood. This work demonstrates that the in vitro association of the two main components of a membrane, the non-bilayer lipid monogalactosyldiacylglycerol (MGDG) and the chlorophyll-a/b light-harvesting antenna protein of photosystem II (LHCII) of pea thylakoids, leads to the formation of large, ordered lamellar structures: (i) thin-section electron microscopy and circular dichroism spectroscopy reveal that the addition of MGDG induces the transformation of isolated, disordered macroaggregates of LHCII into stacked lamellar aggregates with a long-range chiral order of the complexes; (ii) small-angle x-ray scattering discloses that LHCII perturbs the structure of the pure lipid and destroys the inverted hexagonal phase; and (iii) an analysis of electron micrographs of negatively stained 2D crystals indicates that in MGDG-LHCII the complexes are found in an ordered macroarray. It is proposed that, by limiting the space available for MGDG in the macroaggregate, LHCII inhibits formation of the inverted hexagonal phase of lipids; in thylakoids, a spatial limitation is likely to be imposed by the high concentration of membrane-associated proteins.

Circular Dichroism↗

PubMed2000

New evidence for gel-liquid crystalline phase coexistence in the ripple phase of phosphatidylcholines.

Experimental evidence supporting the hypothesis of gel-liquid crystalline phase coexistence in the stable ripple phase of diacylphosphatidylcholines has been obtained from time-resolved X-ray small- (SAXS) and wide-angle diffraction (WAXS) in the millisecond to second time domain. The pretransition of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) exhibits a thin lamellar liquid crystalline intermediate phase (designated Lalpha) if driven far away from equilibrium by an infrared temperature jump (T-jump) technique. The findings can be described by a two-step model. (1) Instantaneously with the T-jump, an anomalously thin lamellar liquid crystalline intermediate phase (d = 5.6-5.8 nm) forms, coexisting with the original gel-phase Lbeta'. Within the first seconds, the lamellar repeat distance of the intermediate increases to a value of about 6.7 nm. A closer examination of these kinetics reveals two relaxation components: a fast process, proceeding within tenths of a second, and a slow process, on the time scale of a few seconds. (2) Finally, both the liquid crystalline and the gel-phase relax into the stable ripple phase Pbeta'. The total process time of the transition is nearly independent of the addition of NaCl, but varies strongly with the chain length of the lecithin species.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed2000

Self-regulation of the lipid content of membranes by non-bilayer lipids: a hypothesis.

Many biological membranes contain lipids that do not form a lamellar phase but the roles of these lipids are not well understood. An artificial membrane assembled from the main non-bilayer lipid and the major integral protein of pea thylakoids revealed that the protein spatially inhibits the formation of non-bilayer structures in the lamellae. Without this inhibition, excess lipids are secreted, creating lipid reservoirs for metabolism and/or later uptake. This determines the protein:lipid ratio in the membrane and hence the balance between structural flexibility and the stability of the key constituents that participate in cooperative interactions.

Lipid Bilayers↗

PubMed2000

Structural information from multilamellar liposomes at full hydration: full q-range fitting with high quality x-ray data.

We present a method for analyzing small angle x-ray scattering data on multilamellar phospholipid bilayer systems at full hydration. The method utilizes a modified Caillé theory structure factor in combination with a Gaussian model representation of the electron density profile such that it accounts also for the diffuse scattering between Bragg peaks. Thus the method can retrieve structural information even if only a few orders of diffraction are observed. We further introduce a procedure to derive fundamental parameters, such as area per lipid, membrane thickness, and number of water molecules per lipid, directly from the electron density profile without the need of additional volumetric measurements. The theoretical apparatus is applied to experimental data on 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine, and 1, 2-dipalmitoyl-sn-glycero-3-phosphoethanolamine liposome preparations.

Liposomes↗

PubMed1999

Crystal structure of human beta2-glycoprotein I: implications for phospholipid binding and the antiphospholipid syndrome.

The high affinity of human plasma beta2-glycoprotein I (beta(2)GPI), also known as apolipoprotein-H (ApoH), for negatively charged phospholipids determines its implication in a variety of physiological pathways, including blood coagulation and the immune response. beta(2)GPI is considered to be a cofactor for the binding of serum autoantibodies from antiphospholipid syndrome (APS) and correlated with thrombosis, lupus erythematosus and recurrent fetal loss. We solved the beta(2)GPI structure from a crystal form with 84% solvent and present a model containing all 326 amino acid residues and four glycans. The structure reveals four complement control protein modules and a distinctly folding fifth C-terminal domain arranged like beads on a string to form an elongated J-shaped molecule. Domain V folds into a central beta-spiral of four antiparallel beta-sheets with two small helices and an extended C-terminal loop region. It carries a distinct positive charge and the sequence motif CKNKEKKC close to the hydrophobic loop composed of residues LAFW (313-316), resulting in an excellent counterpart for interactions with negatively charged amphiphilic substances. The beta(2)GPI structure reveals potential autoantibody-binding sites and supports mutagenesis studies where Trp316 and CKNKEKKC have been found to be essential for the phospholipid-binding capacity of beta(2)GPI.

Amino Acid Sequence↗

PubMed1999

Microphase separation in low density lipoproteins. Evidence for a fluid triglyceride core below the lipid melting transition.

The structural organization of the neutral lipid core in human low density lipoproteins (LDL) was investigated in physicochemically defined, distinct human LDL subspecies in the density range of 1. 0244-1.0435 g/ml by evaluation of the core lipid transition temperature, chemical composition, and the behavior of spin-labeled core lipids. Calorimetric studies were performed on more than 60 LDL preparations, and the transition temperature, which varied between 19 and 32 degreesC, was correlated to the chemical composition and revealed a discontinuity at a critical cholesteryl ester to triglyceride ratio of approximately 7:1. For electron spin resonance studies, several LDL preparations were probed with spin-labeled cholesteryl esters and triglycerides, respectively. In LDL with a high triglyceride content, both labels exhibited similar mobility behavior. In contrast, in LDL with only small concentrations of triglycerides, the behavior of labeled cholesteryl esters and labeled triglycerides differed distinctly. The cholesteryl esters were strongly immobilized below the transition temperature, whereas the triglycerides remained fluid throughout the measured temperatures. These results suggest that the critical cholesteryl ester to triglyceride mass ratio of 7:1 corresponds to two concentric compartments with a radial ratio of 2:1, where the liquid triglycerides occupy the core, and the cholesteryl esters form the frozen shell. At higher triglyceride contents, the triglyceride molecules insert into the cholesteryl ester shell and depress the peak transition temperature of the LDL core, whereas at lower triglyceride contents, excess cholesteryl esters are dissolved in the core.

Calorimetry, Differential Scanning↗

PubMed1998

Crystallization and preliminary X-ray crystallographic studies on apolipoprotein H (beta2-glycoprotein-I) from human plasma.

Apolipoprotein-H (Apo-H, Mw approximately 50 kDa) is a carbohydrate-rich human-plasma protein which exists in blood serum in the free form as well as distributed between several classes of lipoproteins. Single crystals of apo-H have been obtained and crystallographic data sets have been collected. The crystals belong to the orthorhombic space group C2221, with cell dimensions a = 158. 47, b = 169.25, c = 113.28 A (at 100 K). The data indicate that the crystallographic asymmetric unit contains one tetramer of the protein.

Crystallization↗

PubMed1998

Lalpha-phase separation in phosphatidylcholine-water systems induced by alkali chlorides.

The effects of alkali chlorides on phosphatidylcholine-water bilayer systems in the Lalpha-phase were investigated by using small- and wide-angle X-ray scattering. The ternary system LiCl-POPC-H2O under isothermal conditions has shown that above Li+/POPC molar ratios of 0.1 and a lipid concentration above 5% (w/w), a splitting of the lamellar Bragg diffraction peaks into discrete components indicates a phase separation into different lamellar liquid crystalline (smectic A) phases. It is also shown that in saturated distearoyl phosphatidylcholine and in egg phosphatidylcholine, alkali chlorides induce Lalpha-phase separation. The number and repeat distance of the coexisting lamellar phases depend on the nature and concentration of the alkali chloride, the concentration of the phosphatidylcholine, and the degree of the acyl chain unsaturation.

Chemical Phenomena↗

PubMed1998

High-throughput asymmetric double-crystal monochromator of the SAXS beamline at ELETTRA.

A new high-flux wiggler beamline for fast time-resolved small-angle X-ray scattering (SAXS) based on double-focusing optics has recently commenced operation at the 2 GeV third-generation storage ring ELETTRA at Trieste, Italy. Its non-dispersive double-crystal monochromator contains three pairs of interchangeable asymmetrically cut flat Si(111)-crystal pairs, each of which is optimized for high throughput at one of the three fixed energies 5.4, 8 and 16 keV. To cope with the severe thermal power load produced by a 57-pole wiggler on the first crystal of each pair (up to 5.4 W mm(-2) and 700 W under normal incidence, for 400 mA), grazing angles of 2 degrees and optimized back-cooling have been chosen. This solution allows simultaneously a gain of 2.5-3.0 in throughput and, accordingly, in flux density. Finite-element analysis as well as commissioning tests showed that the cooling layout functions very satisfactorily, and that up to 5 x 10(12) photons s(-1) are available at the sample (at 8 keV and 250 mA), as predicted.

Journal Article↗

PubMed1998

First performance assessment of the small-angle X-ray scattering beamline at ELETTRA.

The double-focusing high-flux wiggler beamline dedicated to small-angle X-ray scattering (SAXS) and wide-angle X-ray scattering (WAXS) at ELETTRA has gone into user operation recently. It has been designed specifically for time-resolved studies of non-crystalline and fibrous materials in the submillisecond time scale, and has been optimized for small-angle scattering measurements. An overview of the beamline status and of some representative results, highlighting the performance of the SAXS beamline, are given.

Journal Article↗

PubMed1998

Thermal stability of apolipoprotein B100 in low-density lipoprotein is disrupted at early stages of oxidation while neutral lipid core organization is conserved.

The time course of the unfolding characteristics of the protein moiety and of the thermotropic behavior of the core-located apolar lipids of highly homogeneous low-density lipoprotein (LDL) subspecies (d 1.030-1.040 g/mL) have been evaluated during transition metal- and azo radical-induced oxidation using differential scanning calorimetry. Apolipoprotein B100 (apo-B100) structure was highly sensitive to oxidative modification; indeed, a significant loss of thermal stability was observed at initial stages irrespective of whether oxidation was mediated by site-specific binding of copper ions or by free radicals generated during decomposition of azo compounds. Subsequently, thermal protein integrity was destroyed, as a result of potentially irreversible protein unfolding, cross-linking reactions, and aggregation. Our results suggest that even minimal oxidative modification of apo-B100 has a major impact on the stability of this large monomeric protein. By contrast, the core lipids, which consist primarily of cholesteryl esters and triglycerides and play a determinant role in the thermal transition occurring near physiological temperature, preserved features of an ordered arrangement even during propagation of lipid peroxidation.

Amidines↗

PubMed1998

Surface Roughness Created by Acidic Dissolution of Synthetic Goethite Monitored with SAXS and N2-Adsorption Isotherms

The development of surface roughness of two synthetic goethites with a high, 176 m2/g, and a low, 7 m2/g, initial specific surface area by acid attack is described in terms of fractal geometry. The dissolution kinetics were influenced by the presence of micropores and small mesopores created during dissolution and leading to an initial increase in surface area causing S-shaped dissolution curves. The fractal dimensions were calculated from the adsorption isotherms obtained with N2 as adsorbate. These values of surface fractal dimension agree quite well with the values of the mass and surface fractal dimension obtained with small angle X-ray scattering (SAXS). Copyright 1998 Academic Press. Copyright 1998Academic Press

Journal Article↗