Synchrotron Small-angle X-ray Diffraction Research Articles

Molecular dynamics of cyclically contracting insect flight muscle in vivo

Flight in insects--which constitute the largest group of species in the animal kingdom--is powered by specialized muscles located within the thorax. In most insects each contraction is triggered not by a motor neuron spike but by mechanical stretch imposed by antagonistic muscles. Whereas 'stretch activation' and its reciprocal phenomenon 'shortening deactivation' are observed to varying extents in all striated muscles, both are particularly prominent in the indirect flight muscles of insects. Here we show changes in thick-filament structure and actin-myosin interactions in living, flying Drosophila with the use of synchrotron small-angle X-ray diffraction. To elicit stable flight behaviour and permit the capture of images at specific phases within the 5-ms wingbeat cycle, we tethered flies within a visual flight simulator. We recorded images of 340 micros duration every 625 micros to create an eight-frame diffraction movie, with each frame reflecting the instantaneous structure of the contractile apparatus. These time-resolved measurements of molecular-level structure provide new insight into the unique ability of insect flight muscle to generate elevated power at high frequency.

The structure of DNA–DLPC–cationic gemini surfactant aggregates: a small angle synchrotron X-ray diffraction study

The structure of aggregates formed by interaction of DNA with unilamellar dilauroylphosphatidylcholine (DLPC) vesicles (DNA:DLPC=1:1 base/mol) in the presence of gemini surfactant butane-1,4-diyl-bis(dodecyldimethylammonium bromide) (C12GS) was investigated using synchrotron small angle X-ray diffraction. In the concentration range C12GS +:DLPC≤1 mol/mol, a condensed lamellar L α c phase was found with a repeat period of lipid bilayer stacking in the range d∼5.70–6.53 nm and the DNA interhelical distance d DNA∼3.52–3.99 nm, depending on the concentration of C12GS. At molar ratio C12GS +:DLPC≥0.35:1, the diffractograms have shown the presence of a second lamellar phase with the repeat period d∼5.31 nm which slightly decreases with increasing concentration of C12GS +. The increasing fraction of this phase in the aggregates with increasing concentration of C12GS supports the association of this phase with microscopic domains enriched by surfactant molecules. The temperature behaviour of aggregates was investigated in the range 25–60 °C and the transversal thermal expansivities of the observed phases were determined.

Glycolipid based cubic nanoparticles: preparation and structural aspects

Kinetically stable cubic colloidal particle dispersion was produced from a glycolipid using a novel preparation strategy based on the dialysis principle. The use of synchrotron small-angle X-ray diffraction (SSAXD) permitted the identification of exact structure of these dispersed particles in the colloidal state. Dynamic light scattering methods were used to obtain size and size distributions. A glycoside, 1- O-phytanyl-β- d-xyloside (β-XP), that exhibits Pn3 m cubic phase in an excess aqueous medium, was used as the lipid material. The dialysis technique includes controlled stirring action both inside and outside of the dialysis membrane tube. Initially, a mixed micellar system composed of β-XP, n-octyl-β- d-glucopyranoside (β-OG) and a triblock copolymer, Pluronic F127 (PL) was prepared in the aqueous medium. About 10 wt.% of PL to lipid weight was found to be sufficient to produce stable colloidal dispersions. The mean volume diameter of these colloidal particles was found to be in the range of 0.85±0.05 μm. The cubic phase structure of these colloidal particles is greatly depended on the final β-OG concentration level in the system. Coexistence of Im3 m and Pn3 m cubic structures has been identified in these colloidal particles. This coexistence has the characteristics of Bonnet relation, which forms a compelling case for the infinite periodic minimal surface (IPMS) descriptions. These colloidal particles could restore pure Pn3 m phase structure, but a longer dialysis time was needed. This work, in general, will open up new possibilities for membrane protein reconstitution and other relevant biological applications using colloidal cubic lipid particles.

Destruction of long-range order recorded with in situ small-angle x-ray diffraction in drying colloidal crystals.

High-resolution synchrotron small-angle x-ray diffraction is applied to characterize the structure and long-range order in a sedimentary hard-sphere colloidal crystal before and during its drying. The principles of the technique and the influence of the coherence properties of the x-ray beam are discussed in detail. The capillary forces generated during the drying process are shown to destroy the long-range order and to break the crystal into smaller crystallites with slightly different orientations. The diffraction is shown to switch from the dynamic regime in the long-range-ordered crystal to nearly kinematic diffraction in the mosaic (short-range-ordered) crystal.

Influence of local anesthetics on the phosphatidylcholine model membrane: small-angle synchrotron X-ray diffraction and neutron scattering study

The phase preferences of egg yolk phosphatidylcholine (EYPC) have been examined in the presence of tertiary amine anesthetics [2-(propyloxy)phenyl]-2-(1-piperidinyl)ethyl ester of carbamic acid (C3A) and [2-(heptyloxy)phenyl]-2-(1-piperidinyl)ethyl ester of carbamic acid (C7A, heptacaine). Using the synchrotron small-angle X-ray diffraction (SAXD), it is shown that the C3A anesthetic induces the cubic and hexagonal ( H I) phases at 2≥C3A:EYPC>0.5 and H 2O:EYPC≤40 molar ratios. In contrast, longer alkyloxy chain homolog C7A has no effect on the bilayer arrangement of EYPC at C7A:EYPC≤1 molar ratios as observed by SAXD in C7A+EYPC mixtures hydrated at H 2O:EYPC≤40 molar ratios, as well as in sonicated C7A+EYPC mixtures hydrated in excess water as proved by the small-angle neutron scattering (SANS). The bilayer thickness d L decreases and the bilayer C7A surface area S C7A increases with the increase of C7A:EYPC molar ratio. It is suggested that the ability of tertiary amine local anesthetics to influence the d L and S C7A values and EYPC polymorphism is caused by their effective molecular shape and by charge. The possibility that anesthetic molecules may exert some of their biological effects by virtue of these properties is discussed.

Depletion-induced colloidal crystals at a wall characterised by small-angle X-ray diffraction

Silica photonic colloidal crystals on a flat glass substrate have been grown using the depletion attraction induced by nonadsorbing polymers. The dried samples have been characterised with high-resolution synchrotron small-angle X-ray diffraction and their surface topography has been studied by atomic force microscopy. The presence of long-range ordered crystals with a thickness as large as 90 planes is demonstrated. However, on some places the films are polycrystalline and contain disordered regions.

Bragg rods and multiple X-ray scattering in random-stacking colloidal crystals.

Synchrotron small-angle x-ray diffraction images of random-stacking-induced Bragg scattering rods are obtained in a wide range of wave vectors from a single colloidal crystal. The results reveal a strong multiple scattering effect, which leads to new features in the diffraction pattern-secondary Bragg rods. We argue that dynamic x-ray diffraction is rather common for high-quality colloidal photonic crystals and should be taken into account.

High-resolution small-angle x-ray diffraction study of long-range order in hard-sphere colloidal crystals.

The long-range order parameters in single crystals of hard colloidal spheres grown in sediments of colloid-polymer mixtures are determined using synchrotron small-angle x-ray diffraction with a resolution of 10(-6) of the wave vector. The interplanar positional order derived from the width of lattice reflections extends over at least 500 lattice planes. The lattice planes are orientationally correlated within approximately 0.1 degrees throughout the crystals, whereas the stacking of hexagonal planes remains random.

Fabrication and Characterization of Large Macroporous Photonic Crystals in Titania

We made large, highly ordered structures consisting of crystals of macropores in titania (TiO2), by template-assisted growth. The crystals were characterized by synchrotron small-angle X-ray diffraction, X-ray absorption, wide-angle X-ray diffraction, scanning electron microscopy, optical microscopy, optical reflectivity, and Raman spectroscopy. Care was taken to make well-ordered templates, by slowly growing colloidal crystals from lightly charged polystyrene latex particles and carefully drying them to form opals. Solid material was deposited in the opal template by precipitation from an alkoxide hydrolysis. Subsequently, the samples were heated to 450 °C to form anatase TiO2 and to remove the latex template, which resulted in a macroporous crystal, inverse opal, or air-sphere crystal. The macropores were close-packed and interconnected by windows, and small additional voids were located at interstices between the pores. The macropores were arranged on a face-centered cubic lattice with domains of more ...

Particle excursions in colloidal crystals.

We have performed synchrotron small-angle x-ray diffraction experiments on charge-stabilized colloidal crystals. The mean squared displacements, obtained from Debye-Waller factors, decrease with increasing density beyond freezing. Irrespective of the range of the repulsion, our data and previous results lie on a common curve, linearly scaled between the melting and close-packed densities. Surprisingly, the excursions are smaller than predicted by theory.

Differences between the pressure- and temperature-induced denaturation and aggregation of beta-lactoglobulin A, B, and AB monitored by FT-IR spectroscopy and small-angle X-ray scattering.

We examined the temperature- and pressure-induced unfolding and aggregation of beta-lactoglobulin (beta-Lg) and its genetic variants A and B up to temperatures of 90 degrees C in the pressure range from 1 bar to 10 kbar. To achieve information simultaneously on the secondary, tertiary, and quaternary structures, we have applied Synchrotron small-angle X-ray diffraction and Fourier transform infrared spectroscopy. Upon heating a beta-Lg solution at pH 7.0, the radius of gyration Rg first decreases, indicating a partial dissociation of the dimer into the monomers, the secondary structures remaining essentially unchanged. Above 50 degrees C, the infrared spectroscopy data reveal a decrease in intramolecular beta-sheet and alpha-helical structures, whereas the contribution of disordered structures increases. Within the temperature range from 50 to 60 degrees C, the appearance of the pair distance distribution function is not altered significantly, whereas the amount of defined secondary structures declines approximately by 10%. Above 60 degrees C the aggregation process of 1% beta-Lg solutions is clearly detectable by the increase in Rg and intermolecular beta-sheet content. The irreversible aggregation is due to intermolecular S-H/S-S interchange reactions and hydrophobic interactions. Upon pressurization at room temperature, the equilibrium between monomers and dimers is also shifted and dissociation of dimers is induced. At pressures of approximately 1300 bar, the amount of beta-sheet and alpha-helical structures decreases and the content of disordered structures increases, indicating the beginning unfolding of the protein which enables aggregation. Contrary to the thermal denaturation process, intermolecular beta-sheet formation is of less importance in pressure-induced protein aggregation and gelation. The spatial extent of the resulting protein clusters is time- and concentration-dependent. The aggregation of a 1% (w/w) solution of A, B, and the mixture AB results in the formation of at least octameric units as can be deduced from the radius of gyration of about 36 A. No differences in the pressure stability of the different genetic variants of beta-Lg are detectable in our FT-IR and SAXS experiments. Even application of higher pressures (up to 10 kbar) does not result in complete unfolding of all beta-Lg variants.

Supramolecular structure of lipopolysaccharide and free lipid A under physiological conditions as determined by synchrotron small-angle X-ray diffraction.

Lipopolysaccharides, the major amphiphilic components of the outer leaflet of the outer membrane of Gram-negative bacteria, may assume various three-dimensional supramolecular structures depending on molecular properties (e.g. chemical structure) and on ambient conditions (e.g. temperature, concentration of divalent cations). We applied synchrotron small-angle X-ray diffraction to investigate the supramolecular structures of natural and synthetic Escherichia-coli-type lipid A, of lipid A from Salmonella minnesota, and of rough mutant lipopolysaccharides of E. coli and S. minnesota under physiological water content (greater than 90%) at different temperatures (20, 37, and 55 degrees C) and at different lipid/divalent cation molar ratios (20:1 to 1:1). We found that in the absence of divalent cations rough mutant lipopolysaccharide and free lipid A form unilamellar structures with the main reflections centered around 4.50 nm for free lipid A, 4.80 nm for Re lipopolysaccharide, and 5.90 nm for Rd1 lipopolysaccharide at 20 degrees C, i.e. below the beta----alpha acyl-chain-melting transition temperature. Above this temperature, the reflections are shifted to 4.30 nm for free lipid A (at 55 degrees C), 4.60 nm for Re lipopolysaccharide (at 37 degrees C), and to 5.50 nm for Rd1 lipopolysaccharide (at 37 degrees C). The addition of divalent cations leads (at lower concentrations, i.e. lipid/cation molar ratios 20:1 to 5:1) to sharper reflections expressing a higher state of order and to a shift of the center of the main reflections lying now at 5.10 nm for free lipid A, 6.40 nm for Re and 7.20 nm for Rd1 lipopolysaccharide at 20 degrees C. At higher concentrations of divalent cations (e.g. lipid/cation molar ratio 1:1), an increasing tendency to form nonlamellar, inverted cubic structures is observed which is indicated by the occurrence of another main periodicity and/or of reflections with spacing ratios 1: square root of 2, 1: square root of 3 of the main periodicity. The tendency to assume inverted cubic structures is only weakly pronounced for rough mutant lipopolysaccharides but dominant for free lipid A even at physiological temperature and divalent cation concentration.