Reversible Assembly of β-Sheet Nanocrystals within Caddisfly Silk

Abstract

Nuclear magnetic resonance (NMR) and X-ray diffraction (XRD) experiments reveal the structural importance of divalent cation–phosphate complexes in the formation of β-sheet nanocrystals from phosphorylated serine-rich regions within aquatic silk from caddisfly larvae of the species Hesperophyla consimilis. Wide angle XRD data on native caddisfly silk show that the silk contains a significant crystalline component with a repetitive orthorhombic unit cell aligned along the fiber axis with dimensions of 5.9 Å × 23.2 Å × 17.3 Å. These nanocrystalline domains depend on multivalent cations, which can be removed through chelation with ethylenediaminetetraacetic acid (EDTA). A comparison of wide angle X-ray diffraction data before and after EDTA treatment reveals that the integrated peak area of reflections corresponding to the nanocrystalline regions decreases by 15–25% while that of the amorphous background reflections increases by 20%, indicating a partial loss of crystallinity. 31P solid-state NMR data on native caddisfly silk also show that the phosphorylated serine-rich motifs transform from a rigid environment to one that is highly mobile and water-solvated after treatment with EDTA. The removal of divalent cations through exchange and chelation has therefore caused a collapse of the β-sheet structure. However, NMR results show that the rigid phosphorus environment is mostly recovered after the silk is re-treated with calcium. The 31P spin–lattice (T1) relaxation times were measured at 7.6 ± 3.1 and 1 ± 0.5 s for this calcium-recovered sample and the native silk sample, respectively. The shorter 31P T1 relaxation times measured for the native silk sample are attributed to the presence of paramagnetic iron that is stripped away during EDTA chelation treatment and replaced with diamagnetic calcium.

Connecting caddisworm silk structure and mechanical properties: combined infrared spectroscopy and mechanical analysis

“…The proposal that H-fibroin (pSX) n motifs form a network of Ca 2+ -stabilized β-domains as the principal toughening mechanism of caddisworm silk is supported by multiple lines of experimental evidence. First, infrared (IR) and NMR spectroscopy, as well as X-ray scattering studies, showed decreased β-structure when Ca 2+ was exchanged with Na + [ 7 , 11 , 12 ]. Second, Ca 2+ -depleted silk fibres behaved mechanically like weak elastomers, with the initial stiffness decreased to 1% of native silk fibres, with no yield-like strain softening and no strain cycle hysteresis [ 7 ].…”

Structural and wetting properties of nature's finest silks (order Embioptera)

“…In this study, we expanded the protein–core model to include a description of how water interacts with the protein core in webspinner silk in the absence of organic solvent and show that exposure to water alone results in a change in macroscopic silk structure and wetting properties. Alternatively, exposure to water may remove metals that stabilize the β-sheet structure, as found in caddisfly silk, and when these metal cations are removed, water can penetrate rigid serine-phosphate regions and solvate the protein structure [ 28 ]. This hypothesis will be probed in future studies.…”

Cross‐Aldol Reaction of Isatin with Acetone Catalyzed by Leucinol: A Mechanistic Investigation

“…The ampholytic character of caddisfly H‐fibroin leads to the formation of complex coacervates during the silk formation stage141 (discussed in Section 7). The negatively charged phosphates are proposed to interact with Ca 2+ ions creating intra‐ and intermolecular bridges that promote subsequent folding into rigid domains similar to β‐crystalline regions in spider and silkworm silks 142. 143 Accordingly, the presence of Ca 2+ has been shown to play a crucial role in the structural and macroscopic mechanical properties of these silk fibers 144.…”