Noncontact mode atomic force microscopy was used to investigate native silk proteins prepared in different ways. Low protein concentrations revealed that single protein molecules exhibit a simple, round shape with apparent diameters of 20-25 nm. Shearing the native protein solutions after extraction from the gland and prior to drying led to a beads-on-a-string assembly at the nanometer scale. Protein concentration had a significant effect on the morphology of the protein assemblies. At higher protein concentrations, shear-induced alignment into nanofibrils was observed, while lower concentrations lead to the formation of much thinner fibrils with a width of about 8 nm.
Silks spun by spiders and insects are independently evolved proteinaceous biomaterials with fascinating physical properties attracting scientists from a wide range of disciplines using a wide range of analytical tools. Here we demonstrate the suitability of neutron small angle scattering (SANS) to investigate the morphology and structure of native silk fibroin at near in vivo conditions. Comparing native and reconstituted silk we observed significant differences in sizes, molecular weights, refolding and interactions. These observations question the validity of a presently widespread approach in silk analysis i.e. studying reconstituted silk with the goal to gain important insights into the mechanisms involved in the formation (storage and spinning) of native silks.
Using atomic force microscopy, we present the first molecular-scale comparison of two of the most important silk dopes, native (NSF) and reconstituted (RSF) silkworm fibroin. We found that both systems depended on shear to show self-assembly. Significant differences in the nature of self-assembly between NSF and RSF were shown. In the highest studied concentration of 1000 mg/L, NSF exhibited assembly into 20-30 nm-wide nanofibrils closely resembling the surface structures found in natural silk fibers. RSF, in contrast, showed no self-assembly whatsoever at the same concentration, which suggests that the reconstitution process significantly disrupts silk's inherent self-assembly capability. At lower concentrations, both RSF and NSF formed fibrils under shear, apparently denatured by the substrate. Using image analysis, we quantified the properties of these self-assembled fibrils as a function of concentration and found low-concentration fibrils of NSF to form larger continuous structures than those of RSF, further supporting NSF's superior self-assembly capabilities.
The imaging beamline (IBL/P05) operated by Helmholtz Zentrum Geesthacht (HZG) at the DESY PETRA III storage ring consists of two experimental stations: A micro tomography and a nano tomography end station.Here an overview of the experimental setups and the data acquisition will be given. In addition some first results out of the wide range of applications using the micro tomography station at P05 will be shown. Furthermore, we present first results of the nano tomography end station. These were obtained with an x-ray microscopy setup, which currently operates at energies of 17.4 and 30 keV using polymer compound refractive lenses (CRLs) and rolled prism lenses. Taken together these results clearly show the high potential of the newly built imaging beamline IBL.
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