Morphological and Structural Aspects of the Extremely Halophilic Archaeon Haloquadratum walsbyi

Ultrathin square cell Haloquadratum walsbyi from the Archaea domain are the most abundant microorganisms in the hypersaline water of coastal salterns and continental salt lakes. In this work, we explore the cell surface of these microorganisms using amplitude-modulation atomic-force microscopy in nearly physiological conditions. We demonstrate the presence of a regular corrugation with a periodicity of 16–20 nm attributed to the surface layer (S-layer) protein lattice, striped domains asymmetrically distributed on the cell faces and peculiar bulges correlated with the presence of intracellular granules. Besides, subsequent images of cell evolution during the drying process indicate the presence of an external capsule that might correspond to the giant protein halomucin, predicted by the genome but never before observed by other microscopy studies

Aggregation of Aß(25-35) on DOPC and DOPC/DHA bilayers: an atomic force microscopy study

β amyloid peptide plays an important role in both the manifestation and progression of Alzheimer disease. It has a tendency to aggregate, forming low-molecular weight soluble oligomers, higher-molecular weight protofibrillar oligomers and insoluble fibrils. The relative importance of these single oligomeric-polymeric species, in relation to the morbidity of the disease, is currently being debated. Here we present an Atomic Force Microscopy (AFM) study of Aβ(25-35) aggregation on hydrophobic dioleoylphosphatidylcholine (DOPC) and DOPC/docosahexaenoic 22∶6 acid (DHA) lipid bilayers. Aβ(25-35) is the smallest fragment retaining the biological activity of the full-length peptide, whereas DOPC and DOPC/DHA lipid bilayers were selected as models of cell-membrane environments characterized by different fluidity. Our results provide evidence that in hydrophobic DOPC and DOPC/DHA lipid bilayers, Aβ(25-35) forms layered aggregates composed of mainly annular structures. The mutual interaction between annular structures and lipid surfaces end-results into a membrane solubilization. The presence of DHA as a membrane-fluidizing agent is essential to protect the membrane from damage caused by interactions with peptide aggregates; to reduces the bilayer defects where the delipidation process starts

Aggregation of Aβ(25-35) on mica.

<p>Topographic AFM images of Aβ(25-35) aggregates grown on mica. Images were acquired in tapping mode and in PBS. (a) After 6 hours, Aβ(25-35) forms a smooth homogeneous layer without any well-formed aggregates (3×3 µm², 2459×2459px²). (b) On a smaller scale the peptide tends to organize into a texture with some discernible protrusions (white ellipse) (500×500 nm², 1228×1228px²).</p

Aβ(25-35) high resolution analysis.

<p>Topographic AFM images (a, c) and corresponding phase images (b, d) performed in PBS on the same area (360×360 nm², 1024×1024 px²) of Aβ(25-35) LA domains on the DOPC bilayer. Images were acquired with an average tip-sample force of 170pN by scanning from left to right (a, b) and from right to left (c, d). Topographies clearly show both globular aggregates (circled) and annular structures (squared). The complex LA aggregate (arrow) in the bottom right-hand corner and the globular aggregate in the top right-hand corner (insets) are perturbed by the tip movement. (e, f) Height (black) and phase (red) line profiles of globular (e) and annular structures (f) measured along the white dashed line in Fig. 6a and 6c. (g,h) Height images of a highly dense globular structure region (286×286 nm², 574×574px²) acquired with two different tip-sample forces. At 220 pN (g), the globular structures are not perturbed, while at 234 pN (h) they are mechanically removed leaving the underlying annular structures. (i, l) AFM height images (770×770 nm², 633×633 px²) of LA on two different regions of the DOPC/DHA bilayer. After 1 h45 min of peptide deposition (i) the LA presents a linear organization highlighted by grey fibres. After 2 h20 min (l) the LA forms a structured layer where linear organization is less visible though still distinguishable. In some locations, it is organized into annular structures (red circles) similar in dimension but more sporadic and of different nature compared to the ones observed on DOPC.</p

DOPC and DOPC/DHA lipid bilayers.

<p>(a) Height AFM images of DOPC (2.5×2.5 µm² 1024×1024px²) and DOPC/DHA (b) (2.5×2.5 512×512px²) lipid bilayers performed in tapping mode and in PBS. The bilayers cover 92% (DOPC) and 96% (DOPC/DHA) of the mica surface. On DOPC lipid particles collected at the border or within defects are due to incomplete bilayer formation or to defective rinsing.</p

Aβ(25-35) aggregation on DOPC/DHA lipid bilayer.

<p>(a-c) 5×2.2 µm² (1024×431px²) topographic AFM images of Aβ(25-35) aggregation on DOPC/DHA lipid bilayers. (d) Graph showing the increasing surface area covered by LA (blue line) and the lipid bilayer evolution over time (red line). LA structures were grown within the first 30 min of peptide deposition while the lipid surface area decreased slowly from 96% to 91%. Data were qualitatively fitted by rational and sigmoidal functions for the DOPC/DHA bilayer (Θ_DOPC/DHA) and the LA domains (Θ_P), respectively. Fitting curves act as guide-to-the-eye. (e) Height distribution histograms measured at t = 0 min (black), 30 min (red), 1 h (green), 1 h45 min (blue), 2 h (cyan) and 2 h20 min (magenta).</p