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This study investigates the effect of oxidized sucrose (OS) on selected functional properties of cast films from lupin protein isolate (LPI). LPI with a protein content larger than 0.9 g g−1 was obtained by alkaline extraction and isoelectric precipitation from bitter narrow‐leaved lupins (Lupinus angustifolius L.). OS was synthesized by vicinal diol cleavage using sodium periodate and was successfully characterized by Fourier‐transform infrared‐, 1H‐nuclear magnetic resonance spectroscopy, and aldehyde titration. Films were produced from heated (85°C, 30 min), alkaline (pH 10), aqueous solutions of LPI (0.1 mL−1), glycerol (300 mg g−1 of LPI) and OS (25, 50, 75, and 100 mg g−1 of LPI). The effect of covalent crosslinking between OS and protein chains was studied by investigating mechanical properties, moisture content, total soluble matter, water vapor permeability, and protein solubility for all films. LPI films produced with the addition of OS showed increased mechanical performance as the tensile strength was increased from 3.5 up to 9.3 MPa and elongation at break values could be raised from 118% to 176%. Taken together with the facts that these films are thermoformable and show improved wet strength compared with control films, make them promising materials for sustainable packaging and short‐term applications in agriculture and forestry. [ABSTRACT FROM AUTHOR]

Cribellate silks, produced by ancient spiders, are fascinating because they feature a highly sophisticated, 3D hierarchical structure consisting of filaments with different diameters and shapes. Here, the smallest and thinnest constituents of the cribellate silk are investigated: nanofibrils that form a dense mesh that is supported by larger fibers. Analysis of their structure via atomic force and transmission electron microscopies shows that they are flattened fibrils, only ≈5 nm thick — thinner than any other natural spider silk fibrils previously reported. In this work, the first mechanical tensile testing experiments on these fibrils are carried out, which reveals that the fibrils show an outstanding extensibility of at least 1100%, almost twice as much as the most stretchable spider silk previously reported. Based on these extraordinary findings, this work significantly expands the parameter space of materials properties attainable by spider silks and provides further insights into their nanomechanics. [ABSTRACT FROM AUTHOR]

Expandable shape‐morphing hydrogels that ensure prolonged site residence, have tailored mechanical integrity and tunability, are biocompatible to minimize side effects and can release drugs over an extended time remain challenging to achieve. Herein, a new class of enzyme‐triggered bovine serum albumin and polyethylene glycol diacrylate hybrid hydrogels is presented, contributing to advancements in controlled drug‐model release and actuation. These hydrogels combine the intrinsic properties of proteins with the resilience of synthetic polymers, offering a versatile application platform. Central to our research is the trypsin‐induced simultaneous functionality of controlled drug model release and dynamic shape changes under physiological trypsin concentrations (0.01% w/v). These hydrogels display tailored mechanical and physical properties and microstructure, which are crucial for biomedical devices, soft robotics, and tissue engineering applications. Additionally, the hydrogels effectively control the release of fluorescein isothiocyanate, a model drug, indicating their potential for highly targeted drug delivery, particularly in the gastrointestinal tract. The study also highlights the significant effect of shape‐morphing on drug release rates under physiological trypsin concentrations. These findings suggest that enzyme‐responsive hybrid protein‐polymer hydrogel actuators with tailored mechanical and physical properties can enhance the precision of drug delivery in biomedical applications. [ABSTRACT FROM AUTHOR]

Expandable shape‐morphing hydrogels that ensure prolonged site residence, have tailored mechanical integrity and tunability, are biocompatible to minimize side effects and can release drugs over an extended time remain challenging to achieve. Herein, a new class of enzyme‐triggered bovine serum albumin and polyethylene glycol diacrylate hybrid hydrogels is presented, contributing to advancements in controlled drug‐model release and actuation. These hydrogels combine the intrinsic properties of proteins with the resilience of synthetic polymers, offering a versatile application platform. Central to our research is the trypsin‐induced simultaneous functionality of controlled drug model release and dynamic shape changes under physiological trypsin concentrations (0.01% w/v). These hydrogels display tailored mechanical and physical properties and microstructure, which are crucial for biomedical devices, soft robotics, and tissue engineering applications. Additionally, the hydrogels effectively control the release of fluorescein isothiocyanate, a model drug, indicating their potential for highly targeted drug delivery, particularly in the gastrointestinal tract. The study also highlights the significant effect of shape‐morphing on drug release rates under physiological trypsin concentrations. These findings suggest that enzyme‐responsive hybrid protein‐polymer hydrogel actuators with tailored mechanical and physical properties can enhance the precision of drug delivery in biomedical applications.

Despite the significant progress in protein‐based materials, creating a tunable protein‐activated hydrogel lens remains an elusive goal. This study leverages the synergistic relationship between protein structural dynamics and polymer hydrogel engineering to introduce a highly transparent protein–polymer actuator. By incorporating bovine serum albumin into polyethyleneglycol diacrylate hydrogels, the authors achieved enhanced light transmittance and conferred actuating capabilities to the hydrogel. Taking advantage of these features, a bilayer protein‐driven hydrogel lens that dynamically modifies its focal length in response to pH changes, mimicking the adaptability of the human lens, is fabricated. The lens demonstrates durability and reproducibility, highlighting its potential for repetitive applications. This integration of protein‐diverse biochemistry, folding nanomechanics, and polymer engineering opens up new avenues for harnessing the wide range of proteins to potentially propel various fields such as diagnostics, lab‐on‐chip, and deep‐tissue bio‐optics, advancing the understanding of incorporating biomaterials in the optical field. [ABSTRACT FROM AUTHOR]

Abstract Despite the significant progress in protein‐based materials, creating a tunable protein‐activated hydrogel lens remains an elusive goal. This study leverages the synergistic relationship between protein structural dynamics and polymer hydrogel engineering to introduce a highly transparent protein–polymer actuator. By incorporating bovine serum albumin into polyethyleneglycol diacrylate hydrogels, the authors achieved enhanced light transmittance and conferred actuating capabilities to the hydrogel. Taking advantage of these features, a bilayer protein‐driven hydrogel lens that dynamically modifies its focal length in response to pH changes, mimicking the adaptability of the human lens, is fabricated. The lens demonstrates durability and reproducibility, highlighting its potential for repetitive applications. This integration of protein‐diverse biochemistry, folding nanomechanics, and polymer engineering opens up new avenues for harnessing the wide range of proteins to potentially propel various fields such as diagnostics, lab‐on‐chip, and deep‐tissue bio‐optics, advancing the understanding of incorporating biomaterials in the optical field.

Developing bioinspired materials to convert sunlight into electricity efficiently is paramount for sustainable energy production. Fluorescent proteins are promising candidates as photoactive materials due to their high fluorescence quantum yield and absorption extinction coefficients in aqueous media. However, developing artificial bioinspired photosynthetic systems requires a detailed understanding of molecular interactions and energy transfer mechanisms in the required operating conditions. Here, the supramolecular self-assembly and photophysical properties of fluorescent proteins complexed with organic dyes are investigated in aqueous media. Supercharged mGreenLantern protein, mutated to have a charge of +22, is complexed together with anionic zinc phthalocyanines having 4 or 16 carboxylate groups. The structural characterization reveals a strong electrostatic interaction between the moieties, accompanied by partial conformational distortion of the protein structure, yet without compromising the mGreenLantern chromophore integrity as suggested by the lack of emission features related to the neutral form of the chromophore. The self-assembled biohybrid shows a total quenching of protein fluorescence, in favor of an energy transfer process from the protein to the phthalocyanine, as demonstrated by fluorescence lifetime and ultrafast transient absorption measurements. These results provide insight into the rich photophysics of fluorescent protein-dye complexes, anticipating their applicability as water-based photoactive materials., (© 2024 The Author(s). Advanced Biology published by Wiley‐VCH GmbH.)

Drug delivery systems comprising drug carriers capable of adhering to intestinal tissue have considerable potential to realize more sophisticated systemic drug delivery and topical drug treatments in the intestinal tract. The development of innovative strategies for improving the adhesion efficiency of carriers is of high importance for the advancement of this field. Herein, a novel approach to achieving high adhesion efficiency of drug carriers is presented, where the accessibility of the carrier to the intestinal surface and its subsequent adhesion to the intestinal tissue are promoted by utilizing the thermodynamic tendency of the hydrophobic carrier and its dispersion solvent, triacetin, to be excluded from the aqueous environment. Drug carriers are fabricated using proteins, imparting multiple functions, including drug release and the removal of reactive oxygen species (ROS). Results of ex vivo studies indicate that this multifunctional protein‐based carrier, "protein micropatch," adheres to various mouse intestinal tissues, including the small intestine, colon, and inflamed colon, with high efficiency. Furthermore, protein micropatches, administered to mice via oral or rectal routes, successfully adhere to the intestinal tract. This approach and the highly functionalized carrier described in the study have the potential to significantly contribute to the development of bioadhesive carrier‐based drug delivery systems. [ABSTRACT FROM AUTHOR]

Stimuli responsive hydrogels that can change shape in response to applied external stimuli are appealing for soft robotics, biomedical devices, drug delivery, and actuators. However, existing 3D printed shape morphing materials are non‐biodegradable, which limits their use in biomedical applications. Here, 3D printed protein‐based hydrogels are developed and applied for programmable structural changes under the action of temperature, pH, or an enzyme. Key to the success of this strategy is the use of methacrylated bovine serum albumin (MA–BSA) as a biodegradable building block to Pickering emulsion gels in the presence of N‐isopropylacrylamide or 2‐dimethylaminoethyl methacrylate. These shear‐thinning gels are ideal for direct ink write (DIW) 3D printing of multi‐layered stimuli‐responsive hydrogels. While poly(N‐isopropylacrylamide) and poly(dimethylaminoethyl methacrylate) introduce temperature and pH‐responsive properties into the printed objects, a unique feature of this strategy is an enzyme‐triggered shape transformation based on the degradation of the bovine serum albumin network. To highlight this technique, protein‐based hydrogels that reversibly change shape based on environmental temperature and pH are fabricated, and irreversibly altered by enzymatic degradation, which demonstrates the complexity that can be introduced into 4D printed systems. [ABSTRACT FROM AUTHOR]

While bio-based but chemically synthesized polymers such as polylactic acid require industrial conditions for biodegradation, protein-based materials are home compostable and show high potential for disposable products that are not collected. However, so far, such materials lack in their mechanical properties to reach the requirements for, e.g., packaging applications. Relevant measures for such a modification of protein-based materials are plasticization and cross-linking; the former increasing the elasticity and the latter the tensile strength of the polymer matrix. The assessment shows that compared to other polymers, the major bottleneck of proteins is their complex structure, which can, if developed accordingly, be used to design materials with desired functional properties. Chemicals can act as cross-linkers but require controlled reaction conditions. Physical methods such as heat curing and radiation show higher effectiveness but are not easy to control and can even damage the polymer backbone. Concerning plasticization, effectiveness and compatibility follow opposite trends due to weak interactions between the plasticizer and the protein. Internal plasticization by covalent bonding surpasses these limitations but requires further research specific for each protein. In addition, synergistic approaches, where different plasticization/cross-linking methods are combined, have shown high potential and emphasize the complexity in the design of the polymer matrix.

Untethered mobile micromachines have considerable potential to realize more effective and minimally invasive medicine. Although diverse medical micromachines have been reported over the past few decades, these machines were developed for performing only specific tasks and the functions imparted to them were limited to a few. Hence, the methodologies for imparting a wide variety of functions to machines have not been fully explored. In this study, a novel construction strategy for the multifunctional micromachines is presented, where a specific function can be added to the machine in one step by directly combining the protein-based component, possessing the biological function of constituent proteins, to an arbitrary position of the machine by using an inkjet printing technique. As a proof-of-concept demonstration, various types of machines were constructed by combining multiple components with different functions. These constructed machines successfully performed functions as diverse as enzyme-powered self-propulsion, collection of target objects, including the bilirubin and living cells, enzyme-mediated conversion of substrate molecules to different ones, magnetic guidance, and release of anti-inflammatory drug diapocynin. The study's progressive approach as well as multifunctional and biocompatible machines composed of proteins will profoundly impact the development of intelligent machines equipped with multiplex sophisticated functionalities.

Introduction: Treatment options for cancer metastases, the primary cause of cancer mortality, are limited. The chemokine receptor CXCR4 is an attractive therapeutic target in cancer because it mediates metastasis by inducing cancer cell and macrophage migration. Here we engineered carrier-free CXCR4-targeting RNA-protein nanoplexes that not only inhibited cellular migration but also polarized macrophages to the M1 phenotype. Materials and Methods: A CXCR4-targeting single-chain variable fragment (scFv) antibody was fused to a 3030 Da RNA-binding protamine peptide (RSQSRSRYYRQRQRSRRRRRRS). Self-assembling nanoplexes were formed by mixing the CXCR4-scFv-protamine fusion protein (CXCR4-scFv-RBM) with miR-127-5p, a miRNA shown to mediate M1 macrophage polarization. RNA-protein nanoplexes were characterized with regard to their physicochemical properties and therapeutic efficacy. Results: CXCR4-targeting RNA-protein nanoplexes simultaneously acted as a targeting ligand, a macrophage polarizing drug, and a miRNA delivery vehicle. Our carrier-free, RNA-protein nanoplexes specifically bound to CXCR4-positive macrophages and breast cancer cells, showed high drug loading (~ 90% w/w), and are non-toxic. Further, these RNA-protein nanoplexes significantly inhibited cancer and immune cell migration (75 to 99%), robustly polarized macrophages to the tumor-suppressive M1 phenotype, and inhibited tumor growth in a mouse model of triple-negative breast cancer Conclusions: We engineered a novel class of non-toxic RNA-protein nanoplexes that modulate the tumor stroma. These nanoplexes are promising candidates for add-ons to clinically approved chemotherapeutics. [ABSTRACT FROM AUTHOR]

Protein-based materials are finding new uses and applications after millennia of impacting the daily life of humans. Some of the earliest uses of protein-based materials are still evident in silk and wool textiles and leather goods. Today, even as silks, wools and leathers are still be used in traditional ways, these proteins are now seen as promising materials for biomaterials, vehicles of drug delivery and components of high-tech fabrics. With the advent of biosynthetic methods and streamlined means of protein purification, protein-based materials—recombinant and otherwise—are being used in a host of applications at the cutting edge of medicine, electronics, materials science and even fashion. This commentary aims to discuss a handful of these applications while taking a critical look at where protein-based materials may be used in the future.

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.

In this study we explored the capacity of a mass spectrometry technique, ToF-SIMS, coupled with an immunological method, Dot-ELISA, to characterize the organic binders used in a 13th century painted wooden panel. The panel, which was in a poor state of conservation with only some residual painting left, was a useful bench specimen for these techniques. ToF-SIMS and Dot-ELISA results both indicated the presence of a mixture of rabbit glue and casein and rule out the use of egg in the painted layer of the samples. The results of this study show the capability of the combined use of the two methods to univocally identify protein-based binders in paintings. [ABSTRACT FROM AUTHOR]

To understand if and how extraction conditions influence properties of molded protein films, Crambe abyssinica protein concentrates were produced from deoiled seed meal under various extraction conditions. Properties of the resulting hot compression molded films were evaluated through the molecular weight distribution, protein polymerization behavior and tensile tests. Precipitated protein concentrates demonstrated higher protein content and a pronounced shift to higher molecular weight distributions and reduced solubility on heating, indicating increased protein polymerization compared to those from lyophilized supernatants. Thermally processed films from isoelectrically precipitated protein concentrates show a high resistance to extraction with a combination of reducing agent and denaturant, indicating the presence of non-disulfide covalent cross linking. Also, tensile strength was higher in concentrates from precipitated proteins compared to those from supernatants. The protein concentrates resulting in thermally processed films with a high protein content, the highest levels of protein-protein interaction and high tensile strength were based on alkaline extraction and isoelectric precipitation. Therefore, a combination of alkali extraction and isoelectric precipitation is recommended to produce protein concentrates for molded film production. [ABSTRACT FROM AUTHOR]

Fertilization is an increasingly common practice in horticulture. Nevertheless, the conventionally used fertilization method is ineffective, and it generates contamination problems due to excess nutrients. Therefore, new technologies, such as nanofertilization or controlled-release systems of fertilizers, are currently being tested. Thus, the main objective of this work was to develop controlled-release systems for micronutrients, using soy protein as raw material. Different micronutrients (zinc, copper, iron, and manganese) were evaluated, as well as their incorporation in the form of micro and nanoparticles. The mechanical and functional properties (water uptake capacity, biodegradability, and micronutrient release) of the systems, as well as their use in crops, were studied to assess their viability. The results showed the great potential of these systems to incorporate micronutrients into crops, especially when combined with nanotechnology, improving the benefits of conventional fertilization. [Display omitted] • Soy protein-based systems allow to release micronutrients in a control way. • Controlled release systems are more effective than conventional fertilization. • Nanoparticles achieve more efficient systems with a high water uptake capacity. • Crops obtained from these systems are larger and richer in micronutrients. [ABSTRACT FROM AUTHOR]

The recombinant protein Ultrabithorax (Ubx), a Drosophila melanogaster Hox transcription factor, self-assembles in vitro into biocompatible materials that are remarkably extensible and strong. Here, it is demonstrated that the strength of Ubx materials is due to intermolecular dityrosine bonds. Ubx materials autofluoresce blue, a characteristic of dityrosine, and bind dityrosine-specific antibodies. Monitoring the fluorescence of reduced Ubx fibers upon oxygen exposure reveals biphasic bond formation kinetics. Two dityrosine bonds in Ubx are identified by site-directed mutagenesis followed by measurements of fiber fluorescence intensity. One bond is located between the N-terminus and the homeodomain (Y4/Y296 or Y12/Y293), and another bond is formed by Y167 and Y240. Fiber fluorescence closely correlates with fiber strength, demonstrating that these bonds are intermolecular. This is the first identification of specific residues that participate in dityrosine bonds in protein-based materials. The percentage of Ubx molecules harboring both bonds can be decreased or increased by mutagenesis, providing an additional mechanism to control the mechanical properties of Ubx materials. Duplication of tyrosine-containing motifs in Ubx increases dityrosine content in Ubx fibers, suggesting these motifs could be inserted in other self-assembling proteins to strengthen the corresponding materials. [ABSTRACT FROM AUTHOR]

Enzyme immobilization is an attractive route for achieving catalytically functional surfaces suitable for both continuous and repeated use. Herein, genetic engineering is used to combine the catalytic ability of a xylanase with the self-assembly properties of recombinant spider silk, realizing silk materials with enzymatic activity. Under near-physiological conditions, soluble xylanase-silk fusion proteins assembled into fibers displaying catalytic activity. Also, a xylanase-silk protein variant with the silk part miniaturized to contain only the C-terminal domain of the silk protein formed fibers with catalytic activity. The repertoire of xylanase-silk formats is further extended to include 2D surface coatings and 3D foams, also being catalytically active, showing the versatile range of possible silk materials. The stability of the xylanase-silk materials is explored, demonstrating the possibility of storage, reuse, and cleaning with ethanol. Interestingly, fibers can also be stored dried with substantial residual activity after rehydration. Moreover, a continuous enzymatic reaction using xylanase-silk is demonstrated, making enzymatic batch reactions not the sole possible implementation. The proof-of-concept for recombinantly produced enzyme-silk, herein shown with a xylanase, implies that also other enzymes can be used in similar setups. It is envisioned that the concept of enzyme-silk can find its applicability in, for example, multienzyme reaction systems or biosensors. [ABSTRACT FROM AUTHOR]

Although the in vivo function of the Drosophila melanogaster Hox protein Ultrabithorax (Ubx) is to regulate transcription, in vitro Ubx hierarchically self-assembles to form nanoscale to macroscale materials. The morphology, mechanical properties, and functionality (via protein chimeras) of Ubx materials are all easily engineered. Ubx materials are also compatible with cells in culture. These properties make Ubx attractive as a potential tissue engineering scaffold, but to be used as such they must be biocompatible and nonimmunogenic. In this study, we assess whether Ubx materials are suitable for in vivo applications. When implanted into mice, Ubx fibers attracted few immune cells to the implant area. Sera from mice implanted with Ubx contain little to no antibodies capable of recognizing Ubx. Furthermore, Ubx fibers cultured with macrophages in vitro did not lyse or activate the macrophages, as measured by TNF- α and NO secretion. Finally, Ubx fibers do not cause hemolysis when incubated with human red blood cells. The minimal effects observed are comparable with those induced by biomaterials used successfully in vivo. We conclude Ubx materials are biocompatible and nonimmunogenic. © 2014 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 103A: 1546-1553, 2015. [ABSTRACT FROM AUTHOR]

Proteins implement most of the vital molecular functions of living organisms, including structural support, energy generation, biomolecule sensing, and chemical catalysis, storage, and degradation. While capturing proteins in materials could create devices that mimic these functions, this process is challenging due to the sensitivity of protein structure to the chemical environment. Using recombinant DNA methods, specific functions can be incorporated by fusing the gene encoding a self-assembling protein and the desired functional protein, to produce a single polypeptide that self-assembles into functionalized materials. However, the functional protein has the potential to disrupt protein production, protein assembly, and/or the structure and mechanical properties of the resulting materials. 24 fusion proteins are created based on Ultrabithorax, a Drosophila transcription factor that self-assembles into materials in vitro. The appended proteins dictate the solubility and purification yield of the corresponding protein fusions. Any loss of solubility and yield can be mitigated by fusing a third protein that is highly soluble. All protein fusions self-assemble equally well to produce materials with similar morphologies. Fusing enhanced green fluorescent protein to Ultrabithorax influences mechanical properties of the resulting fibers. It is concluded that a far wider range of proteins can be successfully incorporated into elastomeric protein-based materials than originally anticipated. [ABSTRACT FROM AUTHOR]

The Drosophila melanogaster Hox protein ultrabithorax (Ubx) has the interesting ability to hierarchically self-assemble in vitro into materials that have mechanical properties comparable to natural elastin. Ubx materials can be easily functionalized by gene fusion, generating potentially useful scaffolds for cell and tissue engineering. Here, we tested the cytocompatibility of fibers composed of Ubx or an mCherry-Ubx fusion protein. Fibers were cultured with three primary human cell lines derived from vasculature at low passage: umbilical vein endothelial cells, brain vascular pericytes, or aortic smooth muscle cells. No direct or indirect toxicity was observed for any cell line, in response to fibers composed of either plain Ubx or mCherry-Ubx. Cells readily adhered to Ubx fibers, and cells attached to fibers could be transferred between tissue cultures without loss of viability for at least 96 h. When attached to fibers, the morphology of the three cell lines differed somewhat, but all cells in contact with Ubx fibers exhibited a microtubular network aligned with the long axis of Ubx fibers. Thus, Ubx fibers are cytocompatible with cultured primary human vascular cells. © 2013 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 102A: 97-104, 2014. [ABSTRACT FROM AUTHOR]

Abstract: The choice of biomaterials available for regenerative medicine continues to grow rapidly, with new materials often claiming advantages over the short-comings of those already in existence. Going back to nature, collagen is one of the most abundant proteins in mammals and its role is essential to our way of life. It can therefore be obtained from many sources including porcine, bovine, equine or human and offer a great promise as a biomimetic scaffold for regenerative medicine. Using naturally derived collagen, extracellular matrices (ECMs), as surgical materials have become established practice for a number of years. For clinical use the goal has been to preserve as much of the composition and structure of the ECM as possible without adverse effects to the recipient. This review will therefore cover in-depth both naturally and synthetically produced collagen matrices. Furthermore the production of more sophisticated three dimensional collagen scaffolds that provide cues at nano-, micro‐ and meso-scale for molecules, cells, proteins and bulk fluids by inducing fibrils alignments, embossing and layered configuration through the application of plastic compression technology will be discussed in details. This review will also shed light on both naturally and synthetically derived collagen products that have been available in the market for several purposes including neural repair, as cosmetic for the treatment of dermatologic defects, haemostatic agents, mucosal wound dressing and guided bone regeneration membrane. There are other several potential applications of collagen still under investigations and they are also covered in this review. [Copyright &y& Elsevier]

Untethered mobile micromachines hold great promise in the development of effective and minimally invasive therapies. Although diverse medical micromachines for specific applications have been developed over the past few decades, the coordinated action of multiple machines with different functions remains largely unexplored. In this study, we created three types of biocompatible micromachines using proteins and demonstrated the potential of their coordinated action for medical applications. As a proof of concept, we demonstrated neural replacement therapy, in which neuroblastomas were killed by using an anticancer prodrug and the first machine that contains enzymes, enabling the conversion of the prodrug into a cytotoxic drug. Subsequently, a second machine composed of extracellular matrix was placed on the dead cancer cells to provide a suitable environment for cell adhesion, on which embryonic stem (ES) cells and stromal cells that promote neural differentiation of stem cells were attached by using third machines capable of delivering cells to target positions with desired patterns. As a result, neuroblastomas were replaced with novel healthy neurons derived from ES cells by teaming multiple protein-based machines. We believe that this work highlights the potential of heterogeneous machine groups for medical treatment and the utility of highly biocompatible and functional micromachines made from proteins, representing an important step forward in building more sophisticated micromachine-based therapies.

Elastin-like recombinant protein polymers are a new family of polymers which are captivating the attention of a broad audience ranging from nanotechnologists to biomaterials and more basic scientists. This is due to the extraordinary confluence of different properties shown by this kind of material that are not found together in other polymer systems. Elastin-like polymers are extraordinarily biocompatible, acutely smart and show uncommon self-assembling capabilities. Additionally, they are highly versatile, since these properties can be tuned and expanded in many different ways by substituting the amino acids of the dominating repeating peptide or by inserting, in the polymer architecture, (bio)functional domains extracted from other natural proteins or de novo designs. Recently, the potential shown by elastin-like polymers has, in addition, been boosted and amplified by the use of recombinant DNA technologies. By this means, complex molecular designs and extreme control over the amino-acid sequence can be attained. Nowadays, the degree of complexity and control shown by the elastin-like protein polymers is well beyond the reach of even the most advanced polymer chemistry technologies. This will open new possibilities in obtaining synthetic advanced bio- and nanomaterials. This review explores the present development of elastin-like protein polymers, with a particular emphasis for biomedical uses, along with some future directions that this field will likely explore in the near future. [ABSTRACT FROM AUTHOR]

Nature uses proteins as building blocks to create three-dimensional (3D) structural components (like spiderwebs and tissue) that are recycled within a closed loop. Furthermore, it is difficult to replicate the mechanical properties of these 3D architectures within synthetic systems. In the absence of biological machinery, protein-based materials can be difficult to process and can have a limited range of mechanical properties. Herein, we present an additive manufacturing workflow to fabricate tough, protein-based composite hydrogels and bioplastics with a range of mechanical properties. Briefly, methacrylated bovine-serum-albumin-based aqueous resins were 3D-printed using a commercial vat photopolymerization system. The printed structures were then treated with tannic acid to introduce additional non-covalent interactions and form tough hydrogels. The hydrogel material could be sutured and withstand mechanical load, even after immersion in water for 24 h. Additionally, a denaturing thermal cure could be used to virtually eliminate rehydration of the material and form a bioplastic. To highlight the functionality of this material, a bioplastic screw was 3D-printed and driven into wood without damage to the screw. Moreover, the 3D-printed constructs enzymatically degraded up to 85% after 30 days in pepsin solution. Thus, these protein-based 3D-printed constructs show great potential for biomedical devices that degrade in situ .

Hydrogels made from globular proteins cross-linked covalently into a stable network are becoming an important type of biomaterial, with applications in artificial tissue design and cell culture scaffolds, and represent a promising system to study the mechanical and biochemical unfolding of proteins in crowded environments. Due to the small size of the globular protein domains, typically 2-5 nm, the primary network allows for a limited transfer of protein molecules and prevents the passing of particles and aggregates with dimensions over 100 nm. Here, we demonstrate a method to produce protein materials with micrometer-sized pores and increased permeability. Our approach relies on forming two competing networks: a covalent network made from cross-linked bovine serum albumin (BSA) proteins via a light-activated reaction and a physical network triggered by the aggregation of a polysaccharide, alginate, in the presence of Ca 2+ ions. By fine-tuning the reaction times, we produce porous-protein hydrogels that retain the mechanical characteristics of their less-porous counterparts. We further describe a simple model to investigate the kinetic balance between the nucleation of alginate and cross-linking of BSA molecules and find the upper rate of the alginate aggregation reaction driving pore formation. By enabling a more significant permeability for protein-based materials without compromising their mechanical response, our method opens new vistas into studying protein-protein interactions and cell growth and designing novel affinity methods.

While bio-based but chemically synthesized polymers such as polylactic acid require industrial conditions for biodegradation, protein-based materials are home compostable and show high potential for disposable products that are not collected. However, so far, such materials lack in their mechanical properties to reach the requirements for, e.g., packaging applications. Relevant measures for such a modification of protein-based materials are plasticization and cross-linking; the former increasing the elasticity and the latter the tensile strength of the polymer matrix. The assessment shows that compared to other polymers, the major bottleneck of proteins is their complex structure, which can, if developed accordingly, be used to design materials with desired functional properties. Chemicals can act as cross-linkers but require controlled reaction conditions. Physical methods such as heat curing and radiation show higher effectiveness but are not easy to control and can even damage the polymer backbone. Concerning plasticization, effectiveness and compatibility follow opposite trends due to weak interactions between the plasticizer and the protein. Internal plasticization by covalent bonding surpasses these limitations but requires further research specific for each protein. In addition, synergistic approaches, where different plasticization/cross-linking methods are combined, have shown high potential and emphasize the complexity in the design of the polymer matrix.

The billion tons of synthetic-polymer-based materials (i.e. plastics) produced yearly are a great challenge for humanity. Nature produces even more natural polymers, yet they are sustainable. Proteins are sequence-defined natural polymers that are constantly recycled when living systems feed. Digestion is the protein depolymerization into amino acids (the monomers) followed by their re-assembly into new proteins of arbitrarily different sequence and function. This breaks a common recycling paradigm where a material is recycled into itself. Organisms feed off of random protein mixtures that are "recycled" into new proteins whose identity depends on the cell's specific needs. In this study, mixtures of several peptides and/or proteins are depolymerized into their amino acid constituents, and these amino acids are used to synthesize new fluorescent, and bioactive proteins extracellularly by using an amino-acid-free, cell-free transcription-translation (TX-TL) system. Specifically, three peptides (magainin II, glucagon, and somatostatin 28) are digested using thermolysin first and then using leucine aminopeptidase. The amino acids so produced are added to a commercial TX-TL system to produce fluorescent proteins. Furthermore, proteins with high relevance in materials engineering (β-lactoglobulin films, used for water filtration, or silk fibroin solutions) are successfully recycled into biotechnologically relevant proteins (fluorescent proteins, catechol 2,3-dioxygenase)., (© 2021 The Authors. Advanced Materials published by Wiley-VCH GmbH.)

Lead sulfide (PbS) has drawn research interest due to its excellent optical, thermoelectric, and other physical properties. Various methods have been developed to synthesize PbS nanoparticles (NPs) for application in a variety of fields. However, most of these methods either require complex experimental conditions or are inefficient. Here, a highly efficient approach to synthesize PbS NPs, inspired by using 2,3-dimercaptosuccinic acid (DMSA) as a heavy metal chelator to treat lead, mercury, and arsenic poisoning, is reported. DMSA can attach to protein-based materials, such as human hair, using disulfide bonds (–SS–) as reaction sites. DMSA tethered to the protein-based materials can further chelate with Pb2+ to assist in the formation of PbS NPs. Characterization of the synthesized PbS NPs demonstrated clear crystallization and uniform distribution. Furthermore, the new method is much more efficient than those previously reported, which can greatly improve the production of PbS NPs for a broad range of applications. Moreover, detailed analysis shows that all protein-based materials can function as templates for synthesizing PbS NPs due to the prevalent existence of –SS– in the natural materials. [ABSTRACT FROM AUTHOR]

Microbially produced protein-based materials (PBMs) are appealing due to use of renewable feedstock, low energy requirements, tunable side-chain chemistry, and biodegradability. However, high-strength PBMs typically have high molecular weights (HMW) and repetitive sequences that are difficult to microbially produce due to genetic instability and metabolic burden. We report the development of a biosynthetic strategy termed seeded chain-growth polymerization (SCP) for synthesis of HMW PBMs in living bacterial cells. SCP uses split intein (SI) chemistry to cotranslationally polymerize relatively small, genetically stable material protein subunits, effectively preventing intramolecular cyclization. We apply SCP to bioproduction of spider silk in Escherichia coli , generating HMW spider silk proteins (spidroins) up to 300 kDa, resulting in spidroin fibers of high strength, modulus, and toughness. SCP provides a modular strategy to synthesize HMW, repetitive material proteins, and may facilitate bioproduction of a variety of high-performance PBMs for broad applications.

Protein-based materials are finding new uses and applications after millennia of impacting the daily life of humans. Some of the earliest uses of protein-based materials are still evident in silk and wool textiles and leather goods. Today, even as silks, wools and leathers are still be used in traditional ways, these proteins are now seen as promising materials for biomaterials, vehicles of drug delivery and components of high-tech fabrics. With the advent of biosynthetic methods and streamlined means of protein purification, protein-based materials-recombinant and otherwise-are being used in a host of applications at the cutting edge of medicine, electronics, materials science and even fashion. This commentary aims to discuss a handful of these applications while taking a critical look at where protein-based materials may be used in the future., (© 2018 The Authors.)

As interest in using proteins to assemble functional, biocompatible, and environmentally friendly materials is growing, developing scalable protocols for producing recombinant proteins with customized functions coupled to straightforward fabrication processes is becoming crucial. Here, we use E. coli bacteria to produce amyloid protein nanofibers that are key constituents of the biofilm extracellular matrix and show that protein nanofiber aggregates can be purified using a fast and easily accessible vacuum filtration procedure. With their extreme resistance to heat, detergents, solvents, and denaturing agents, engineered curli nanofibers remain functional throughout the rigorous processing and can be used to assemble macroscopic materials directly from broth culture. As a demonstration, we show that engineered curli nanofibers can be fabricated into self-standing films while maintaining the functionality of various fused domains that confer new specific binding activity to the material. We also demonstrate that purified curli fibers can be disassembled, reassembled into thin films, and recycled for further materials processing. Our scalable approach, which combines established purification techniques for amyloid fibers, is applicable to a new class of recombinant amyloid proteins whose sequence can be easily tailored for diverse applications through genetic engineering.

Although the in vivo function of the Drosophila melanogaster Hox protein Ultrabithorax (Ubx) is to regulate transcription, in vitro Ubx hierarchically self-assembles to form nanoscale to macroscale materials. The morphology, mechanical properties, and functionality (via protein chimeras) of Ubx materials are all easily engineered. Ubx materials are also compatible with cells in culture. These properties make Ubx attractive as a potential tissue engineering scaffold, but to be used as such they must be biocompatible and nonimmunogenic. In this study, we assess whether Ubx materials are suitable for in vivo applications. When implanted into mice, Ubx fibers attracted few immune cells to the implant area. Sera from mice implanted with Ubx contain little to no antibodies capable of recognizing Ubx. Furthermore, Ubx fibers cultured with macrophages in vitro did not lyse or activate the macrophages, as measured by TNF-α and NO secretion. Finally, Ubx fibers do not cause hemolysis when incubated with human red blood cells. The minimal effects observed are comparable with those induced by biomaterials used successfully in vivo. We conclude Ubx materials are biocompatible and nonimmunogenic., (© 2014 Wiley Periodicals, Inc.)