Spider Silk Research Articles

Preparation of Strong and Thermally Conductive, Spider Silk-Inspired, Soybean Protein-Based Adhesive for Thermally Conductive Wood-Based Composites.

The development of formaldehyde-free functional wood composite materials through the preparation of strong and multifunctional soybean protein adhesives to replace formaldehyde-based resins is an important research area. However, ensuring the bonding performance of soybean protein adhesive while simultaneously developing thermally conductive adhesive and its corresponding wood composites is challenging. Taking inspiration from the microphase separation structure of spider silk, boron nitride (BN) and soy protein isolate (SPI) were mixed by ball milling to obtain a BN@SPI matrix and combined with the self-synthesized hyperbranched reactive substrates as amorphous region reinforcer and cross-linker triglycidylamine to prepare strong and thermally conductive soybean protein adhesive with cross-linked microphase separation structure. These findings indicate that mechanical ball milling can be employed to strip BN followed by combination with SPI, resulting in a tight bonded interface connection. Subsequently, the adhesive's dry and wet shear strengths increased by 14.3% and 90.5% to 1.83 and 1.05 MPa, respectively. The resultant adhesive also possesses a good thermal conductivity (0.363 W/mK). Impressively, because hot-pressing helps the resultant adhesive to establish a thermal conduction pathway, the thermal conductivity of the resulting wood-based composite is 10 times higher than that of the SPI adhesive, which shows a thermal conductivity similar to that of ceramic tile and has excellent potential for developing biothermal conductivity materials, geothermal floors, and energy storage materials. Moreover, the adhesive possessed effective flame retardancy (limit oxygen index = 36.5%) and mildew resistance (>50 days). This bionic design represents an efficient technique for developing multifunctional biomass adhesives and composites.

Designed Multifunctional Spider Silk Enabled by Genetically Encoded Click Chemistry (Adv. Funct. Mater. 43/2023)

Designed Multifunctional Spider Silk Enabled by Genetically Encoded Click Chemistry (Adv. Funct. Mater. 43/2023)

A bioinspired synthetic fused protein adhesive from barnacle cement and spider dragline for potential biomedical materials

Biomaterials with excellent biocompatibility, mechanical performance, and self-recovery properties are urgently needed for tissue regeneration. Inspired by barnacle cement and spider silk, we genetically designed and overexpressed a fused protein (cp19k-MaSp1) composed of Megabalanus rosa (cp19k) and Nephila clavata dragline silk protein (MaSp1) in Pichia pastoris. The recombinant cp19k-MaSp1 exhibited enhanced adhesion capability beyond those of the individual proteins in both aqueous and non-aqueous conditions. cp19k-MaSp1 protein fiber scaffolds prepared through electrospinning have adequate hydrophilicity compared to cp19k and MaSp1 protein fiber scaffolds, and offer improved overall porosity compared to MaSp1 protein fiber scaffolds. The cp19k-MaSp1 protein fiber scaffolds showed excellent proteolytically stable properties because of only 9.6 % depletion after incubation in a biodegradation solution for 56 d. The cp19k-MaSp1 protein fiber scaffolds present remarkably high extreme tensile strength (112.7 ± 11.6 MPa) and superior ductility (438.4 ± 43.9 %) compared with cp19k (34.4 ± 8.1 MPa, 115.4 ± 32.7 %) and MaSp1 protein fiber scaffolds (65.8 ± 9.3 MPa, 409.6 ± 23.1 %), also 68.4 % of tensile strength was recovered by incubation in K+ buffer after multiple stretches, which create a favorable cell adhesion, growth, and proliferation environment for human umbilical vein endothelial cells (HUVECs). The improved biocompatibility, extensive adhesion, mechanical strength, and self-recovery properties make the bioinspired synthetic cp19k-MaSp1 a potential candidate for biomedical tissue reconstruction.

Could an Anterior Cruciate Ligament Be Tissue-Engineered from Silk?

Silk has a long history as an exclusive textile, but also as a suture thread in medicine; nowadays, diverse cell carriers are manufactured from silk. Its advantages are manifold, including high biocompatibility, biomechanical strength and processability (approved for nearly all manufacturing techniques). Silk's limitations, such as scarcity and batch to batch variations, are overcome by gene technology, which allows for the upscaled production of recombinant "designed" silk proteins. For processing thin fibroin filaments, the sericin component is generally removed (degumming). In contrast to many synthetic biomaterials, fibroin allows for superior cell adherence and growth. In addition, silk grafts demonstrate superior mechanical performance and long-term stability, making them attractive for anterior cruciate ligament (ACL) tissue engineering. Looking at these promising properties, this review focusses on the responses of cell types to silk variants, as well as their biomechanical properties, which are relevant for ACL tissue engineering. Meanwhile, sericin has also attracted increasing interest and has been proposed as a bioactive biomaterial with antimicrobial properties. But so far, fibroin was exclusively used for experimental ACL tissue engineering approaches, and fibroin from spider silk also seems not to have been applied. To improve the bone integration of ACL grafts, silk scaffolds with osteogenic functionalization, silk-based tunnel fillers and interference screws have been developed. Nevertheless, signaling pathways stimulated by silk components remain barely elucidated, but need to be considered during the development of optimized silk cell carriers for ACL tissue engineering.

Highly strong and tough silk by feeding silkworms with rare earth ion-modified diets

Nature-derived silk fibers possess excellent biocompatibility, sustainability, and mechanical properties, yet producing strong and tough silk fibers in a facile and large-scale manner remains a significant challenge. Herein, we report a simple method for preparing strong and tough silk fibers by feeding silkworms rare earth ion-modified diets. The resulting silk fibers exhibit significantly increased tensile strength and toughness, with average values of 0.85 ± 0.07 GPa and 156 ± 13 MJ m−3, respectively, and maximum values of 0.97 ± 0.04 GPa and 188 ± 19 MJ m−3, approaching those of spider dragline silk. Our findings suggest that the incorporation of rare earth ions (La3+ or Eu3+) into the silk fibers contributes to this enhancement. Structure analysis reveals a reduction in content and an improvement in orientation of β-sheet nanocrystals in silk fibers. X-ray photoelectron spectroscopy analysis confirms the chemical interaction between rare earth ions with β-sheet nanocrystals. The structural evolution and chemical interactions lead to the simultaneous enhancement in both strength and toughness. This work presents a simple, scalable, and effective strategy for producing ultra-strong and tough silk fibers with potential applications in areas requiring super structural materials, such as personal protection and aerospace.

UV-curing polyurethane pressure-sensitive adhesive with high shear strength and good adhesion properties inspired by spider silk

It is a challenge to develop polyurethane pressure-sensitive adhesive (PU-PSA) simultaneously with good cohesion and adhesion properties. Inspired by the structure of multiple hydrogen bonds in spider silk, we introduced Acylsemicarbazide (ASCZ) moiety that can form multiple hydrogen bonds in the polymer network into PU-PSA. The characteristic absorption peaks of ester groups at 1600–1780 cm−1 were analyzed in FTIR testing, and the results showed that ASCZ formed multiple hydrogen bonds in PU-PSA, and the hydrogen bond content increased with the increase of ASCZ content. In particular, PUA-AD20% with 24.3 % hydrogen bond content showed higher shear strength (0.51 MPa) and debonding energy (3.3 KJ m−2). Rheological tests showed that PU-PSA storage modulus and loss modulus increased with increasing frequency, which was highly frequency dependent. Therefore, as dynamic and reversible sacrificial bonds, hydrogen bonds can dissipate energy efficiently and resist external forces. In addition, the ASCZ component also provided PU-PSA with the polar groups required for adhesion. Therefore, PUA-AD20% had excellent adhesion properties. Compared to PUA, PUA-AD20% tack and peel strength increased by 36.1 % and 146.8 %, with values of 4.9 N and 11.6 N, respectively. In summary, this work offered a simple and versatile approach to designing PU-PSA with outstanding overall performance.

High strength, high stiffness and toughness, defect‐tolerant waterborne polyurethane with healing and antibacterial abilities

AbstractThe nature creates many biomaterials such as spider silk which exhibits a combination of stiffness, strength and toughness. However, most of synthetic unfilled materials suffer from a trade‐off between toughness and stiffness. Inspired by the structure of spider silk but beyond it, we proposed a novel molecular design to achieve transparent unfilled waterborne polyurethane (WPU) with simultaneously enhanced stiffness (280.9 MPa), tensile strength (25.1 MPa) and toughness (140.0 MJ/m3) as well as good elasticity (710%). The designed WPU comprised homogeneous continuous phase (soft segments) and diverse H‐bonds (hard segments) dispersed in it. The increase of rigid molecular chain content and H‐bonds contributed to the high stiffness of WPU. Furthermore, the mismatch of stiffness between hard domains and soft segments might promote crack deflection and branching, which endowed the robust WPU with fracture energy of 81.16 kJ/m2. The robust WPU film could be healed to recover most of its original mechanical properties (strength for 24.4 MPa and elongation for 610%) under heating. In addition, the WPU films demonstrated good antibacterial performance against Staphylococcus aureus and Escherichia coli after chlorination.

High-strength and ultra-tough whole spider silk fibers spun from transgenic silkworms

High-strength and ultra-tough whole spider silk fibers spun from transgenic silkworms

Research Progress on Spider‐Inspired Tough Fibers†

Comprehensive SummarySpider silk has attracted increasing attention due to its fascinating combination of ultra‐high tenacity high strength, and excellent elasticity. Based on the fundamental biological studies on spider silk, significant research efforts have been devoted to biotechnology and chemical synthesis to mimic or even exceed the properties of natural spider silk fibers. Moreover, the natural spider silk fiber has been simulated with the burgeoning development of numerous spinning technologies, including wet spinning, dry spinning, electrostatic spinning, and microfluidic spinning, which continuously help to optimize the properties of synthetic spider silk. The unique characteristics of natural spider silk include high refraction transmission, heat resistance, antimicrobial properties, biocompatibility, and super shrinking. Biconical recreation of spider silk with special features and extraordinary capabilities demonstrates potential applications in biomedicine, smart wearables, artificial muscles and sensors, aerospace and other domains.

A Minispidroin Guides the Molecular Design for Cellular Condensation Mechanisms in S. cerevisiae.

Structural engineering of molecules for condensation is an emerging technique within synthetic biology. Liquid-liquid phase separation of biomolecules leading to condensation is a central step in the assembly of biological materials into their functional forms. Intracellular condensates can also function within cells in a regulatory manner to facilitate reaction pathways and to compartmentalize interactions. We need to develop a strong understanding of how to design molecules for condensates and how their in vivo-in vitro properties are related. The spider silk protein NT2RepCT undergoes condensation during its fiber-forming process. Using parallel in vivo and in vitro characterization, in this study, we mapped the effects of intracellular conditions for NT2RepCT and its several structural variants. We found that intracellular conditions may suppress to some extent condensation whereas molecular crowding affects both condensate properties and their formation. Intracellular characterization of protein condensation allowed experiments on pH effects and solubilization to be performed within yeast cells. The growth of intracellular NT2RepCT condensates was restricted, and Ostwald ripening was not observed in yeast cells, in contrast to earlier observations in E. coli. Our results lead the way to using intracellular condensation to screen for properties of molecular assembly. For characterizing different structural variants, intracellular functional characterization can eliminate the need for time-consuming batch purification and in vitro condensation. Therefore, we suggest that the in vivo-in vitro understanding will become useful in, e.g., high-throughput screening for molecular functions and in strategies for designing tunable intracellular condensates.

NMR assignment and dynamics of the dimeric form of soluble C-terminal domain major ampullate spidroin 2 from Latrodectus hesperus.

Spider dragline silk has attracted great interest due to its outstanding mechanical properties, which exceed those of man-made synthetic materials. Dragline silk, which is composed of at least major ampullate spider silk protein 1 and 2 (MaSp1 and MaSp2), contains a long repetitive domain flanked by N-terminal and C-terminal domains (NTD and CTD). Despite the small size of the CTD, this domain plays a crucial role as a molecular switch that regulates and directs spider silk self-assembly. In this study, we report the 1H, 13C, and 15N chemical shift assignments of the Latrodectus hesperus MaSp2 CTD in dimeric form at pH 7. Our solution NMR data demonstrated that this protein contains five helix regions connected by a flexible linker.

Auditory Neurons on the Silk Road – spider silk for bridging the nerve-electrode-gap

Abstract The cochlear implant (CI) restores hearing to patients with severe to profound sensorineural hearing loss by stimulating the spiral ganglion neurons (SGN). Due to the inner ear anatomy, a fluid-filled gap remains between the SGN and the electrode array. This gap impedes focused stimulation and thus optimized performance with the CI. Regenerating neurites may bridge this fluid-filled gap for a direct nerveelectrode- link but require a supportive matrix. Spider silk might be a suitable candidate for this support. Therefore, silk fibers were tested for biocompatibility with inner ear neuronal tissue, suitability as neurite outgrowth matrix and applicability to the CI-electrode array. Dragline silk from female Trichonephila spiders was woven around a frame and autoclaved. Spiral ganglia of young rats were prepared, cut and placed in a drop of medium on the parallel silk fibers for pre-cultivation to support adherence before remaining medium was added. After 5d of cultivation, cells were fixed and immunocytologically stained. For CIapplication, silk was wrapped around a silicone dummy and a one-sided spacer. Opposite the spacer, alginate-hydrogel was applied for silk-fixation and -shielding. After gelation, silk loops were cut in the middle to form protruding threads. Cells showed a high degree of migration and neurite regeneration along the silk fibers and in some cases the outgrown neurites directly contacted the silk. It was possible to fix the silk to the dummy as protruding fibers. The tested spider silk is compatible with the inner ear tissue in culture, supports cell growth and seems to be an attractive material for neurite contacting. Application to the CI as protruding fibers is possible, making it a promising candidate for structural support to bridge the nerve-electrode-gap.

Inspired by spider silk: Ultra-tough and exceptionally stretchable coating

Inspired by spider silk: Ultra-tough and exceptionally stretchable coating

Natural spider silk nanofibrils produced by assembling molecules or disassembling fibers.

Spider silk is biocompatible, biodegradable, and rivals some of the best synthetic materials in terms of strength and toughness. Despite extensive research, comprehensive experimental evidence of the formation and morphology of its internal structure is still limited and controversially discussed. Here, we report the complete mechanical decomposition of natural silk fibers from the golden silk orb-weaver Trichonephila clavipes into ≈10 nm-diameter nanofibrils, the material's apparent fundamental building blocks. Furthermore, we produced nanofibrils of virtually identical morphology by triggering an intrinsic self-assembly mechanism of the silk proteins. Independent physico-chemical fibrillation triggers were revealed, enabling fiber assembly from stored precursors "at-will". This knowledge furthers the understanding of this exceptional material's fundamentals, and ultimately, leads toward the realization of silk-based high-performance materials. STATEMENT OF SIGNIFICANCE: Spider silk is one of the strongest and toughest biomaterials, rivaling the best man-made materials. The origins of these traits are still under debate but are mostly attributed to the material's intriguing hierarchical structure. Here we fully disassembled spider silk into 10 nm-diameter nanofibrils for the first time and showed that nanofibrils of the same appearance can be produced via molecular self-assembly of spider silk proteins under certain conditions. This shows that nanofibrils are the key structural elements in silk and leads toward the production of high-performance future materials inspired by spider silk.

Engineering SilkCatcher/Tag pairs for production of native-sized spider silk proteins

Engineering SilkCatcher/Tag pairs for production of native-sized spider silk proteins

Factors Influencing Properties of Spider Silk Coatings and Their Interactions within a Biological Environment.

Biomaterials are an indispensable part of biomedical research. However, although many materials display suitable application-specific properties, they provide only poor biocompatibility when implanted into a human/animal body leading to inflammation and rejection reactions. Coatings made of spider silk proteins are promising alternatives for various applications since they are biocompatible, non-toxic and anti-inflammatory. Nevertheless, the biological response toward a spider silk coating cannot be generalized. The properties of spider silk coatings are influenced by many factors, including silk source, solvent, the substrate to be coated, pre- and post-treatments and the processing technique. All these factors consequently affect the biological response of the environment and the putative application of the appropriate silk coating. Here, we summarize recently identified factors to be considered before spider silk processing as well as physicochemical characterization methods. Furthermore, we highlight important results of biological evaluations to emphasize the importance of adjustability and adaption to a specific application. Finally, we provide an experimental matrix of parameters to be considered for a specific application and a guided biological response as exemplarily tested with two different fibroblast cell lines.

Spider‐Silk‐Inspired Heterogeneous Supramolecular Network with Strain‐Stiffening, High Damping Capacity, and Supercontraction

AbstractNatural materials are adaptive with intriguing mechanical properties such as strain‐stiffening, high damping, and stimuli‐responsive actuation under ambient conditions, which are evolutionarily derived to adapt to variable environments. Such adaptabilities are highly desirable for advanced smart materials in soft robots and bionic applications yet rarely achieved all in one synthetic material. Inspired by the molecular chemistry and structure of spider silk, a structurally heterogeneous supramolecular network with all‐round adaptabilities is developed by evaporation‐induced self‐assembly of hydrogen‐bonded macromolecules. The supramolecular network consists of both hard (crystallites) and soft (amorphous) phases with dynamic hydrogen bonds, exhibiting strong strain, time, and hydration dependency with adaptive mechanical and structural responses to varying strain, deformation rate as well as moisture. Through multifunctional crosslinking, the network exhibits silk‐like attributes with an extensibility of >130%, intense strain‐stiffening (sixfold modulus enhancement), high damping capacity (>80%) over a wide range of strain rate, and moisture‐triggered large supercontraction (contraction ratio of >50%). Artificial materials with such combined adaptiveness under bio‐benign conditions are promising for applications in biomimetic and biomedical fields.

Self-Assembly of RGD-Functionalized Recombinant Spider Silk Protein into Microspheres in Physiological Buffer and in the Presence of Hyaluronic Acid.

Biomaterials made of self-assembling protein building blocks are widely explored for biomedical applications, for example, as drug carriers, tissue engineering scaffolds, and functionalized coatings. It has previously been shown that a recombinant spider silk protein functionalized with a cell binding motif from fibronectin, FN-4RepCT (FN-silk), self-assembles into fibrillar structures at interfaces, i.e., membranes, fibers, or foams at liquid/air interfaces, and fibrillar coatings at liquid/solid interfaces. Recently, we observed that FN-silk also assembles into microspheres in the bulk of a physiological buffer (PBS) solution. Herein, we investigate the self-assembly process of FN-silk into microspheres in the bulk and how its progression is affected by the presence of hyaluronic acid (HA), both in solution and in a cross-linked HA hydrogel. Moreover, we characterize the size, morphology, mesostructure, and protein secondary structure of the FN-silk microspheres prepared in PBS and HA. Finally, we examine how the FN-silk microspheres can be used to mediate cell adhesion and spreading of human mesenchymal stem cells (hMSCs) during cell culture. These investigations contribute to our fundamental understanding of the self-assembly of silk protein into materials and demonstrate the use of silk microspheres as additives for cell culture applications.

High-strength, self-reinforcing and recyclable multifunctional lignin-based polyurethanes based on multi-level dynamic cross-linking

The preparation of lignin-based polyurethanes with excellent mechanical properties and recyclability remains extremely challenging because of the poor compatibility of lignin and matrix. Inspired by the multiple structures of spider silk and pearl layers, high-strength, heat-resistant and recyclable lignin-based polyurethanes (LPUs) were prepared by introducing weak hydrogen bonds, strong hydrogen bonds and dynamic silyl ether covalent bonds. Multiple hydrogen bonds provided a large energy dissipation mechanism. The covalent linkage between lignin and matrix promoted their compatibility and formed a robust cross-linking network. Depending on the multi-level dynamic interactions, the prepared elastomer exhibited a mechanical strength of 73 MPa, which was 1.9 times higher than that of the blank sample, with an initial thermal decomposition temperature of 310.5 °C and a good recyclability. Especially, the mechanical strength of LPUs was up to 99 MPa after mechanical training. In addition, the abundant conjugated structure of lignin endowed LPUs with an excellent photothermal conversion ability. Under the irradiation of NIR light with 4000 w m−2, the surface temperature increased to 95 °C within two minutes, exhibiting a stable photothermal performance even after 6 cycles. This work showed the application prospects of LPUs in the fields of high strength engineering materials and photo-induced actuators.

Pulling and analyzing silk fibers from aqueous solution using a robotic device

Spiders, silkworms, and many other animals can spin silk with exceptional properties. However, artificially spun fibers often fall short of their natural counterparts partly due sub-optimal production methods. A variety of methods, such as wet-, dry-, and biomimetic spinning have been used. The methods are based on extrusion, whereas natural spinning also involves pulling. Another shortcoming is that there is a lack feedback control during extension. Here we demonstrate a robotic fiber pulling device that enables controlled pulling of silk fibers and in situ measurement of extensional forces during the pulling and tensile testing of the pulled fibers. The pulling device was used to study two types of silk—one recombinant spider silk (a structural variant of ADF3) and one regenerated silk fibroin. Also, dextran—a branched polysaccharide—was used as a reference material for the procedure due to its straightforward preparation and storage. No post-treatments were applied. The pulled regenerated silk fibroin fibers achieved high tensile strength in comparison to similar extrusion-based methods. The mechanical properties of the recombinant spider silk fibers seemed to be affected by the liquid-liquid phase separation of the silk proteins.