Squid Ring Teeth Research Articles

Engineering advanced cellulosics for enhanced triboelectric performance using biomanufactured proteins

Biological polymers, such as polysaccharides and polypeptides, offer renewable and biodegradable solutions for a more sustainable future. These polymers comprise natural building blocks, such as amino acids and glycans, which ensure their true environmental benefits at the end of their lifecycle. For example, cellulose is a highly sustainable material with many excellent properties, including renewability, biodegradability, and versatility in its functionality. It can be used in various forms, such as textiles, packaging materials, and building insulation. Here, we studied advanced cellulosic materials produced by blending or creating bi-composites with biomanufactured proteins inspired by squid ring teeth (SRT). Biomanufactured proteins can be synthesized in larger quantities, have a controlled production process, be modified to create desirable variants, and their production can be scaled up or down. Specifically, we engineered recombinant SRT proteins to have high electrostatic charge, induce crystallinity, and provide polar hydroxyl groups, which enhances cellulosic materials’ triboelectric response. The triboelectric voltage of blend triacetate and cellulose fibers increased by 72–108% and 49–57%, respectively, with a protein content of 10% wt. Furthermore, coating proteins on cellulosic fibers to create bi-composite fibers is a highly effective method for doubling (200%) the triboelectric performance. This finding has important implications for developing sustainable triboelectric materials and producing advanced materials using biomanufacturing.

Cephalopod inspired self-healing protein foams for oil-water separation

Cephalopods are remarkable creatures, captivating scientists with their advanced neurophysiology, complex behavior, and miraculously effective camouflage. Research into cephalopods has led to many discoveries in neuroscience, cell biology, and materials science. Specifically, squids provide us with remarkable self-healing Squid Ring Teeth protein, which is applied herein to extend the life span of foams. Despite the advantages of porosity in surface science applications, porosity impairs mechanical properties by making materials more prone to structural damage-which traditional polymeric foams also suffer from. Drawing inspiration from Squid Ring Teeth, we developed self-healing tandem repeat proteins to overcome these challenges. By leveraging porosity and self-healing properties inspired by Squid Ring Teeth, we created bioengineered protein foams with high separation capacity (5.1g g-1) and efficiency (≈94%). The foams healed entirely within minutes which regained over 100% strength after repair. These advances promise applications for efficient continuous water treatment through durable filter cartridges.

Bioinspired stretchable molecular composites of 2D-layered materials and tandem repeat proteins

Protein based composites, such as nacre and bone, show astounding evolutionary capabilities, including tunable physical properties. Inspired by natural composites, we studied assembly of atomistically thin inorganic sheets with genetically engineered polymeric proteins to achieve mechanically compliant and ultra-tough materials. Although bare inorganic nanosheets are brittle, we designed flexible composites with proteins, which are insensitive to flaws due to critical structural length scale (∼2 nm). These proteins, inspired by squid ring teeth, adhere to inorganic sheets via secondary structures (i.e., β-sheets and α-helices), which is essential for producing high stretchability (59 ± 1% fracture strain) and toughness (54.8 ± 2 MJ/m3). We find that the mechanical properties can be optimized by adjusting the protein molecular weight and tandem repetition. These exceptional mechanical responses greatly exceed the current state-of-the-art stretchability for layered composites by over a factor of three, demonstrating the promise of engineering materials with reconfigurable physical properties.

Diffusive Dynamic Modes of Recombinant Squid Ring Teeth Proteins by Neutron Spectroscopy.

Stimuli-responsive structural proteins are emerging as promising biocompatible materials for a wide range of biological and nonbiological applications. To understand the physical properties of structural proteins and to replicate their performance in biosynthetic systems, there is a need to understand the molecular mechanisms and relationships that regulate their structure, dynamics, and properties. Here, we study the dynamics of a recombinant squid-inspired protein from Loligo vulgaris (Lv18) by elastic and quasielastic neutron scattering (QENS) to understand the connection between nanostructure, chain dynamics, and mechanical properties. Lv18 is a semicrystalline structural protein, which is plasticized by water above its glass transition temperature at 35 °C. Elastic scans revealed an increased protein chain mobility upon hydration, superimposed dynamic processes, and a decrease in dynamic transition temperatures. Further analysis by QENS revealed that while dry Lv18 protein dynamics are dominated by localized methyl group rotations, hydrated Lv18 dynamics are dominated by the confined diffusion of flexible chains within a β-sheet nanocrystalline network (8 Å of confinement radius). Our findings establish a relationship between the segment block architecture of Lv18, the diffusive motions within the protein structure, and the mechanical properties of recombinant squid proteins, which will advance the molecular design of novel high-performance protein-inspired materials with tailored dynamics and mechanical properties.

Enhancing sustainability and elasticity of synthetic fibers by tandem repeat proteins

Protein fiber production in heterologous organisms, such as bacteria, provides a new possibility for engineering high-performance materials and composites. The discovery and design of sustainable materials that are biological or inspired by biological principles are essential for the development and production of the next generation of circular bioeconomy. Here, we created a hybrid of biological and synthetic materials by combining bio-engineered proteins with synthetic acrylic polymers to enhance the sustainability and elasticity of the blend fibers. First, we developed an optimized expression (i.e. yield exceeding 1 g l−1) and purification method (>80% purity) for squid ring teeth inspired by tandem proteins at the facility scale. We showed that our protein-based powder, produced via industrial fermentation, can be manufactured into braided yarns with acrylic using wet-spinning. Our fibers have enhanced elasticity when hydrated due to the hydrogen network between the protein and acrylic fibers.

Identification of new silk-like protein from B. magister and development of functional materials based on it

Tandem repeat proteins have composite structure and unique properties, which allow them to be used in multiple fields, such as soft photonics, drug delivery and textile industry. The recent discovery of squid ring teeth (SRT) proteins have expanded the existing repertoire of repetitive polypeptides. We chose previously unexplored squid B. magister for our research, isolated and analyzed a new protein forming its ring teeth and hooks, and amplified the corresponding gene. Finally, we used this new isolated SRT protein to fabricate transparent thin films and microspheres.

Dielectrophoretic separation of randomly shaped protein particles

Recently, insoluble protein particles have been increasingly investigated for artificial drug delivery systems due to their favorable properties, including programmability for active drug targeting of diseases as well as their biocompatibility and biodegradability after administration. One of the biggest challenges is selectively collecting desirable self-repairable particles in the spherical morphology with monodispersity to enable consistent levels and rates of drug loading and release. Therefore, technology that allows sorting of protein particles with respect to size and morphology will enhance the design and production of next-generation drug delivery materials. Here, we introduce a dielectrophoretic (DEP) separation technique to selectively isolate spherical protein particles from a mixture of randomly shaped particles. We tested this approach by applying it to a mixture of precipitated squid ring teeth inspired tandem repeat protein particles with diverse sizes and morphologies. The DEP trapping system enabled us to isolate specific-sized, spherical protein particles out of this mixture: after separation, the fraction of 2 µm and 4 µm spherical particles was increased from 28.64% of mixture to 80.53% and 74.02% with polydispersity indexes (PDIs) decreased from 0.93 of mixture to 0.19 and 0.09, respectively. The protein particles show high aqueous swelling capability (up to 74% by mass) that could enable delivery of drug solutions. This work is intended to inspire the future development of biocompatible drug-delivery systems.

Highly Conductive Self-Healing Biocomposites Based on Protein Mediated Self-Assembly of PEDOT:PSS Films.

Composites of conducting polymers offer a broad spectrum of materials for interfacing electronic devices with biological systems. Particularly, material systems based on poly(styrenesulfonate) doped poly(3,4-ethylenedioxythiophene) (PEDOT:PSS) have found applications in many bioelectronic devices as biosensitive transistors, controlled drug delivery media, and strain, temperature, and humidity sensors. The biocompatibility, intercoupled electronic and ionic conductivity, and air stable electrical properties render PEDOT:PSS based material systems indispensable for bioelectronics. However, these materials are commonly used in thin film form since freestanding films of pristine PEDOT:PSS are considered mechanically brittle compared to biological tissues, and unlike biological systems these conductive films cannot restore/heal their physical properties after excessive mechanical deformation. Here we report conductive biocomposites of PEDOT:PSS and tandem repeat proteins with the ability to self-heal once plasticized via water. The tandem repeat proteins acquired from squid ring teeth (SRT) induce structural effects on PEDOT:PSS including improved crystallinity and formation of fibrous network structures. These structural effects lead to electrical conductivity values reaching 120 S/cm for biocomposites with SRT protein concentrations below 20 wt %, which exceeds the conductivity of pristine PEDOT:PSS (∼100 S/cm). More importantly, tandem proteins facilitate consistent self-healing of freestanding biocomposites with SRT protein concentrations beyond 40 wt %. These robust biocomposites with high electrical conductivity and the ability to self-heal can find applications in numerous soft electronic systems spanning from implantable, transient, and epidermal electronics to electronic textiles.

Bioinspired and Mechanically Strong Fibers Based on Engineered Non‐Spider Chimeric Proteins

AbstractSilk‐protein‐based fibers have attracted considerable interest due to their low weight and extraordinary mechanical properties. Most studies on fibrous proteins focus on the recombinant spidroins, but these fibers exhibit moderate mechanical performance. Thus, the development of alternative structural proteins for the construction of robust fibers is an attractive goal. Herein, we report a class of biological fibers produced using a designed chimeric protein, which consists of the sequences of a cationic elastin‐like polypeptide and a squid ring teeth protein. Remarkably, the chimeric protein fibers exhibit a breaking strength up to about 630 MPa and a corresponding toughness as high as about 130 MJ m−3, making them superior to many recombinant spider silks and even comparable to some native counterparts. Therefore, this strategy is a novel concept for exploring bioinspired ultrastrong protein materials through the development of new types of structural chimeric proteins.

Bioinspired and Mechanically Strong Fibers Based on Engineered Non-Spider Chimeric Proteins.

Silk-protein-based fibers have attracted considerable interest due to their low weight and extraordinary mechanical properties. Most studies on fibrous proteins focus on the recombinant spidroins, but these fibers exhibit moderate mechanical performance. Thus, the development of alternative structural proteins for the construction of robust fibers is an attractive goal. Herein, we report a class of biological fibers produced using a designed chimeric protein, which consists of the sequences of a cationic elastin-like polypeptide and a squid ring teeth protein. Remarkably, the chimeric protein fibers exhibit a breaking strength up to about 630 MPa and a corresponding toughness as high as about 130 MJ m-3 , making them superior to many recombinant spider silks and even comparable to some native counterparts. Therefore, this strategy is a novel concept for exploring bioinspired ultrastrong protein materials through the development of new types of structural chimeric proteins.

Squid Ring Teeth–coated Mesh Improves Abdominal Wall Repair

Hernia repair is a common surgical procedure with polypropylene (PP) mesh being the standard material for correction because of its durability. However, complications such as seroma and pain are common, and repair failures still approach 15% secondary to poor tissue integration. In an effort to enhance mesh integration, we evaluated the applicability of a squid ring teeth (SRT) protein coating for soft-tissue repair in an abdominal wall defect model. SRT is a biologically derived high-strength protein with strong mechanical properties. We assessed tissue integration, strength, and biocompatibility of a SRT-coated PP mesh in a first-time pilot animal study. PP mesh was coated with SRT (SRT-PP) and tested for mechanical strength against uncoated PP mesh. Cell proliferation and adhesion studies were performed in vitro using a 3T3 cell line. Rats underwent either PP (n = 3) or SRT-PP (n = 6) bridge mesh implantation in an anterior abdominal wall defect model. Repair was assessed clinically and radiographically, with integration evaluated by histology and mechanical testing at 60 days. Cell proliferation was enhanced on SRT-PP mesh. This was corroborated in vivo by abdominal wall histology, dramatically diminished craniocaudal mesh contraction, improved strength testing, and higher tissue failure strain. There was no increase in seroma or visceral adhesion formation. No foreign body reactions were noted on liver histology. SRT applied as a coating appears to augment mesh-tissue integration and improve abdominal wall stability following bridged repair. Further studies in larger animals will determine its applicability for hernia repair in patients.

Inkjet Printing of Self‐Assembled 2D Titanium Carbide and Protein Electrodes for Stimuli‐Responsive Electromagnetic Shielding

Abstract2D titanium carbides (MXene) possess significant characteristics including high conductivity and electromagnetic interference shielding efficiency (EMI SE) that are important for applications in printed and flexible electronics. However, MXene‐based ink formulations are yet to be demonstrated for proper inkjet printing of MXene patterns. Here, tandem repeat synthetic proteins based on squid ring teeth (SRT) are employed as templates of molecular self‐assembly to engineer MXene inks that can be printed as stimuli‐responsive electrodes on various substrates including cellulose paper, glass, and flexible polyethylene terephthalate (PET). MXene electrodes printed on PET substrates are able to display electrical conductivity values as high as 1080 ± 175 S cm−1, which significantly exceeds electrical conductivity values of state‐of‐the‐art inkjet‐printed electrodes composed of other 2D materials including graphene (250 S cm−1) and reduced graphene oxide (340 S cm−1). Furthermore, this high electrical conductivity is sustained under excessive bending deformation. These flexible electrodes also exhibit effective EMI SE values reaching 50 dB at films with thicknesses of 1.35 µm, which mainly originate from their high electrical conductivity and layered structure.

Critical factors for the bulk adhesion of engineered elastomeric proteins.

Many protein-based materials, such as soy and mussel adhesive proteins, have been the subject of scientific and commercial interest. Recently, a variety of protein adhesives have been isolated from diverse sources such as insects, frogs and squid ring teeth. Many of these adhesives have similar amino acid compositions to elastomeric proteins such as elastin. Although elastin is widely investigated for a structural biomaterial, little work has been done to assess its adhesive potential. In this study, recombinant elastin-like polypeptides were created to probe the factors affecting adhesion strength. Lap shear adhesion was used to examine the effects of both extrinsic factors (pH, concentration, cross-linker, humidity, cure time and cure temperature) and intrinsic factors (protein sequence, structure and molecular weight). Of the extrinsic factors tested, only humidity, cure time and cure temperature had a significant effect on adhesion strength. As water content was reduced, adhesion strength increased. Of the intrinsic factors tested, amino acid sequence did not significantly affect adhesion strength, but less protein structure and higher molecular weights increased adhesion strength directly. The strengths of proteins in this study (greater than 2 MPa) were comparable to or higher than those of two commercially available protein-based adhesives, hide glue and a fibrin sealant. These results may provide general rules for the design of adhesives from elastomeric proteins.

Programmable Proton Conduction in Stretchable and Self-Healing Proteins

Proton conduction is ubiquitous in nature and has many applications in energy and electronic technologies. Although protein based materials show bulk proton conduction 10 times lower than conventional ion-conducting materials, they have unique advantages including biocompatibility, self-healing, tunable structure, and fine-grained control of material properties via amino acid sequence. Here, we studied the bulk proton conduction of tandem repeat proteins and demonstrate that tandem repetition of sequences from squid ring teeth (SRT) proteins significantly and systematically enhances bulk proton transport properties. Inelastic neutron scattering experiments between 4 K and 350 K reveal that highly repetitive proteins show enhanced conductivity. Our highly repetitive proteins achieve higher proton conductivity than state-of-the-art biological proton conductors (with peak conductivities of 3.5 mS cm–1), as well as demonstrate unique self-healing characteristics. These proteins also exhibit exceptionally high...

Mechanical Properties of Tandem-Repeat Proteins Are Governed by Network Defects.

Topological defects in highly repetitive structural proteins strongly affect their mechanical properties. However, there are no universal rules for structure-property prediction in structural proteins due to high diversity in their repetitive modules. Here, we studied the mechanical properties of tandem-repeat proteins inspired by squid ring teeth proteins using rheology and tensile experiments as well as spectroscopic and X-ray techniques. We also developed a network model based on entropic elasticity to predict structure-property relationships for these proteins. We demonstrated that shear modulus, elastic modulus, and toughness scale inversely with the number of repeats in these proteins. Through optimization of structural repeats, we obtained highly efficient protein network topologies with 42 MJ/m3 ultimate toughness that are capable of withstanding deformations up to 350% when hydrated. Investigation of topological network defects in structural proteins will improve the prediction of mechanical properties for designing novel protein-based materials.

Research Update: Programmable tandem repeat proteins inspired by squid ring teeth

Cephalopods have evolved many interesting features that can serve as inspiration. Repetitive squid ring teeth (SRT) proteins from cephalopods exhibit properties such as strength, self-healing, and biocompatibility. These proteins have been engineered to design novel adhesives, self-healing textiles, and the assembly of 2d-layered materials. Compared to conventional polymers, repetitive proteins are easy to modify and can assemble in various morphologies and molecular architectures. This research update discusses the molecular biology and materials science of polypeptides inspired by SRT proteins, their properties, and perspectives for future applications.

Structural Protein-Based Whispering Gallery Mode Resonators

Nature provides a set of solutions for photonic structures that are finely tuned, organically diverse, and optically efficient. Exquisite knowledge of structure–property relationships in proteins aids in the design of materials with desired properties for building devices with novel functionalities, which are difficult to achieve or previously unattainable. Here we report whispering-gallery-mode (WGM) microresonators fabricated entirely from semicrystalline structural proteins (i.e., squid ring teeth, SRT, from Loligo vulgaris and its recombinant) with quality factors as high as 105. We first demonstrate versatility of protein-based devices via facile doping, engaging secondary structures. Then we investigate thermorefractivity and find that it increases with β-sheet crystallinity, which can be altered by methanol exposure and is higher in the selected recombinant SRT protein than its native counterpart. We present a set of photonic devices fabricated from SRT proteins such as add-drop filters and fibers....

Programmable molecular composites of tandem proteins with graphene oxide for efficient bimorph actuators

The rapid expansion in the spectrum of two-dimensional (2D) materials has driven research efforts on the fabrication of 2D composites and heterostructures. Highly ordered structure of 2D materials provides an excellent platform for controlling the ultimate structure and properties of the composite material with precision. However, limited control over the structure of the matrix phase and its interactions with highly ordered 2D materials results in defective composites with inferior performance. Here, we demonstrate the successful synthesis, integration, and characterization of hybrid 2D material systems consisting of tandem repeat (TR) proteins inspired by squid ring teeth and graphene oxide (GO). The TR protein layer acts as a unique programmable assembler for GO layers with precise control of interlayer distance of less than 1 nm. As an application, we further demonstrate thermal actuation using bimorph molecular composite films. Bimorph actuators made of molecular composite films (GO/TR) can lead to energy efficiencies 18 times higher than regular bimorph actuators consisting of a GO layer and a TR protein layer (i.e., conventional bulk composite of GO and TR). Additionally, molecular composite bimorph actuators can reach curvature values as high as 1.2 cm−1 by using TR proteins with higher molecular weight, which is 3 times higher than conventional GO and TR composites.

Self-Healing Textile: Enzyme Encapsulated Layer-by-Layer Structural Proteins.

Self-healing materials, which enable an autonomous repair response to damage, are highly desirable for the long-term reliability of woven or nonwoven textiles. Polyelectrolyte layer-by-layer (LbL) films are of considerable interest as self-healing coatings due to the mobility of the components comprising the film. In this work mechanically stable self-healing films were fabricated through construction of a polyelectrolyte LbL film containing squid ring teeth (SRT) proteins. SRTs are structural proteins with unique self-healing properties and high elastic modulus in both dry and wet conditions (>2 GPa) due to their semicrystalline architecture. We demonstrate LbL construction of multilayers containing native and recombinant SRT proteins capable of self-healing defects. Additionally, we show these films are capable of utilizing functional biomolecules by incorporating an enzyme into the SRT multilayer. Urease was chosen as a model enzyme of interest to test its activity via fluorescence assay. Successful construction of the SRT films demonstrates the use of mechanically stable self-healing coatings, which can incorporate biomolecules for more complex protective functionalities for advanced functional fabrics.

Proteins: Materials Fabrication from Native and Recombinant Thermoplastic Squid Proteins (Adv. Funct. Mater. 47/2014)

Squid ring teeth complex extracted from Loligo vulgaris shows reversible solid to melt phase transition on page 7401. M. C. Demirel and team show how the direct extraction or recombinant expression of protein based thermoplastics opens up new avenues for materials fabrication and synthesis, which will eventually be competitive with the high-end synthetic oil based plastics.