Nanoscale Structural Features in Major Ampullate Spider Silk

Spider major ampullate silk is often schematically represented as a two-phase material composed of crystalline nanodomains in an amorphous matrix. Here we are interested in revealing its more complex nanoscale organization by probing <i>Argiope bruennichi</i> dragline-type fibers using scanning X-ray nanodiffraction. This allows resolving transversal structural features such as an about 1 μm skin layer composed of around 100 nm diameter nanofibrils serving presumably as an elastic sheath. The core consists of a composite of several nm size crystalline nanodomains with poly­(l-alanine) microstructure, embedded in a polypeptide network with short-range order. Stacks of nanodomains separated by less ordered nanosegments form nanofibrils with a periodic axial density modulation which is particularly sensitive to radiation damage. The precipitation of larger β-type nanocrystallites in the outer core–shell is attributed to MaSp1 protein molecules

Myelin Organization in the Nodal, Paranodal, and Juxtaparanodal Regions Revealed by Scanning X-Ray Microdiffraction

<div><p>X-ray diffraction has provided extensive information about the arrangement of lipids and proteins in multilamellar myelin. This information has been limited to the abundant inter-nodal regions of the sheath because these regions dominate the scattering when x-ray beams of 100 µm diameter or more are used. Here, we used a 1 µm beam, raster-scanned across a single nerve fiber, to obtain detailed information about the molecular architecture in the nodal, paranodal, and juxtaparanodal regions. Orientation of the lamellar membrane stacks and membrane periodicity varied spatially. In the juxtaparanode-internode, 198–202 Å-period membrane arrays oriented normal to the nerve fiber axis predominated, whereas in the paranode-node, 205–208 Å-period arrays oriented along the fiber direction predominated. In parts of the sheath distal to the node, multiple sets of lamellar reflections were observed at angles to one another, suggesting that the myelin multilayers are deformed at the Schmidt-Lanterman incisures. The calculated electron density of myelin in the different regions exhibited membrane bilayer profiles with varied electron densities at the polar head groups, likely due to different amounts of major myelin proteins (P0 glycoprotein and myelin basic protein). Scattering from the center of the nerve fibers, where the x-rays are incident <i>en face</i> (perpendicular) to the membrane planes, provided information about the lateral distribution of protein. By underscoring the heterogeneity of membrane packing, microdiffraction analysis suggests a powerful new strategy for understanding the underlying molecular foundation of a broad spectrum of myelinopathies dependent on local specializations of myelin structure in both the PNS and CNS.</p></div

Amyloid β Peptide Conformational Changes in the Presence of a Lipid Membrane System

Here we are presenting a comparative analysis of conformational changes of two amyloid β peptides, Aβ(25–35) and Aβ(1–42), in the presence and absence of a phospholipid system, namely, POPC/POPS (1-palmitoyl-2-oleoylphospatidylcholine/palmitoyl-2-oleoylphospatidylserine), through Raman spectroscopy, synchrotron radiation micro Fourier-transform infrared spectroscopy, and micro X-ray diffraction. Ringlike samples were obtained from the evaporation of pure and mixed solutions of the proteins together with the POPC/POPS system on highly hydrophilic substrates. The results confirm the presence of a α-helical to β-sheet transition from the internal rim of the ringlike samples to the external one in the pure Aβ(25–35) residual, probably due to the convective flow inside the droplets sitting on highly hydrophilic substrates enhancing the local concentration of the peptide at the external edge of the dried drop. In contrast, the presence of POPC/POPS lipids in the peptide does not result in α-helical structures and introduces the presence of antiparallel β-sheet material together with parallel β-sheet structures and possible β-turns. As a control, Aβ(1–42) peptide was also tested and shows β-sheet conformations independently from the presence of the lipid system. The μXRD analysis further confirmed these conclusions, showing how the absence of the phospholipid system induces in the Aβ(25–35) a probable composite α/β material while its coexistence with the peptide leads to a not oriented β-sheet conformation. These results open interesting scenarios on the study of conformational changes of Aβ peptides and could help, with further investigations, to better clarify the role of enzymes and alternative lipid systems involved in the amyloidosis process of Aβ fragments

Paranodal-nodal diffraction differs from juxtaparanodal-internodal diffraction.

<p>(<b>A</b>) X-ray diffraction patterns #449 and #481 from the nodal/paranodal region. (<b>B</b>) Intensity distribution (relative scale) as a function of reciprocal coordinate (1/Å), with Bragg orders 2–4 indicated. <i>M</i> and <i>E</i> refer to meridional (red arrow) and equatorial (blue arrow) scatter, respectively. (<b>C</b>) Electron density distribution as a function of distance (Å) from the center of the cytoplasmic apposition. The data for fresh (unfixed) myelin (green curve), which has a period of 176 Å, was obtained as described in <i>Materials and Methods (Data Analysis)</i>.</p

Orientation of patterns reveals changing in wrapping of myelin around axon.

<p>Mapping of the principal orientations of lamellar membranes in the region of a node of Ranvier (<i>arrows</i>) for a single myelinated nerve (<b><i>left</i></b>) and a pair overlapping fibers (<b><i>right</i></b>). To determine the overall orientation of the membrane planes on the nerve fiber, at each position we drew a line parallel to the plane of the membranes and perpendicular to the membrane stacking. The vast majority of reflections in the internode corresponds to membranes oriented parallel to the surface of the fiber. Neighboring the node of Ranvier, however, the lamellar stacks curl around and become more perpendicular to the axis of the nerve fiber (<i>arrows</i>). For clarity, orientation of less intense lamellar scattering is not shown for the single fiber (<i>left panel</i>). On the right, the orientations of the two most intense sets of lamellar reflections are shown by long red and short blue lines, respectively. Detailed analysis in this paper focuses on the image to the right.</p

Position-dependent profiles for lamellar myelin consistent with differing ratios of myelin proteins.

<p>Electron density distribution on an absolute scale for data from diffraction patterns #11E, 18E, and 42E, where <i>E</i> refers to scattering along the equator. The observed and calculated structure factors on a relative scale are indicated in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0100592#pone-0100592-t001" target="_blank">Table 1</a>. (The absolute scaling method is described in <i>S.I., Section 1.</i>) The differences in the levels of electron density in the lipid polar head group layers at the cytoplasmic and extracellular sides of the membrane bilayer are interpreted in terms of a heterogeneous distribution of MBP and P0 glycoprotein in different regions of the myelin sheath (see Text for details).</p

Microdiffraction from single, teased fibers of myelinated axons.

<p>(<b>A</b>) Montage of the small-angle portions of diffraction patterns from a raster scan of a pair of teased nerve fibers. The vertically-oriented nerve has a node of Ranvier slightly above its crossing with the horizontally nerve. Individual frames in (A), circled and numbered, were chosen for detailed analyses described in this paper. A video of stepping through all of the images in (A), with a white dot in the last frame for each row, is available as <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0100592#pone.0100592.s001" target="_blank">Fig. S1</a>. (<b>B</b>) Optical micrograph of the same field of view as the montage. (<b>C</b>) A larger field of view of the nerve fibers.</p

Structural data for characteristic observed diffraction patterns.

<p>The structure factors for the intensities measured by the microfocus beam are given by , and their scaling was determined by , where <i>d</i> is the lamellar period. The diffracting power <i>P</i> was measured according to the equation described in (<b><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0100592#pone.0100592.s005" target="_blank">Text S1</a></b>). The intensity data for fresh, mouse sciatic nerve myelin were obtained from <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0100592#pone-0100592-g005" target="_blank">Figure 5</a> in Ref. (7). In the Table, the sample is indicated by the image number, and the scanning direction is indicated by <i>E</i>, for along the equator (lamellar stacks perpendicular to the vertical oriented nerve fiber; <i>M</i>, for along the meridian (lamellar stacks parallel to the vertically oriented nerve fiber), and <i>25deg</i>, for 25° from the horizontal. The distance between the centers of membrane bilayers across the cytoplasmic apposition is given by <i>2u</i>. <i>Cyt</i>, the width of the cytoplasmic apposition; <i>Lpg</i>, the distance between the lipid polar head group layers; <i>Ext</i>, the width of the extracellular apposition. The number of unit cells <i>N</i>, or coherent length, and lattice disorder Δ were calculated from , where <i>w</i> is the observed integral width for the <i>h^th</i> reflection, and <i>b</i> is the integral width of the direct beam <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0100592#pone.0100592-Inouye3" target="_blank">[54]</a>.</p

Thermal Transformations of Self-Assembled Gold Glyconanoparticles Probed by Combined Nanocalorimetry and X‑ray Nanobeam Scattering

Noble metal nanoparticles with ligand shells are of interest for applications in catalysis, thermo-plasmonics, and others, involving heating processes. To gain insight into the structure-formation processes in such systems, self-assembly of carbohydrate-functionalized gold nanoparticles during precipitation from solution and during further heating to ca. 340 °C was explored by in situ combination of nanobeam SAXS/WAXS and nanocalorimetry. Upon precipitation from solution, X-ray scattering reveals the appearance of small 2D domains of close-packed nanoparticles. During heating, increasing interpenetration of the nanoparticle soft shells in the domains is observed up to ca. 81 °C, followed by cluster formation at ca. 125 °C, which transform into crystalline gold nuclei at around 160 °C. Above ca. 200 °C, one observes the onset of coalescence and grain growth resulting in gold crystallites of average size of about 100 nm. The observed microstructural changes are in agreement with the in situ heat capacity measurements with nanocalorimetry

Directed Growth of Virus Nanofilaments on a Superhydrophobic Surface

The evaporation of single droplets of colloidal tobacco mosaic virus (TMV) nanoparticles on a superhydrophobic surface with a hexagonal pillar-pattern results in the formation of coffee-ring type residues. We imaged surface features by optical, scanning electron, and atomic force microscopies. Bulk features were probed by raster-scan X-ray nanodiffraction. At ∼100 pg/μL nanoparticle concentration, the rim of the residue connects to neighboring pillars via fibrous extensions containing flow-aligned crystalline domains. At ∼1 pg/μL nanoparticle concentration, nanofilaments of ≥80 nm diameter and ∼20 μm length are formed, extending normal to the residue-rim across a range of pillars. X-ray scattering is dominated by the nanofilament form-factor but some evidence for crystallinity has been obtained. The observation of sheets composed of stacks of self-assembled nanoparticles deposited on pillars suggests that the nanofilaments are drawn from a structured droplet interface