Biomimetic repeat protein derived from Xenopus tropicalis for fibrous scaffold fabrication.

Collagen, silk, and elastin are the fibrous proteins consist of representative amino acid repeats. Because these proteins exhibited distinguishing mechanical properties, they have been utilized in diverse applications, such as fiber-based sensors, filtration membranes, supporting materials, and tissue engineering scaffolds. Despite their infinite prevalence and potential, most studies have only focused on a few repeat proteins. In this work, the hypothetical protein with a repeat motif derived from the frog Xenopus tropicalis was obtained and characterized for its potential as a novel protein-based material. The codon-optimized recombinant frog repeat protein, referred to as 'xetro', was produced at a high rate in a bacterial system, and an acid extraction-based purified xetro protein was successfully fabricated into microfibers and nanofibers using wet spinning and electrospinning, respectively. Specifically, the wet-spun xetro microfibers demonstrated about 2- and 1.5-fold higher tensile strength compared with synthetic polymer polylactic acid and cross-linked collagen, respectively. In addition, the wet-spun xetro microfibers showed about sevenfold greater stiffness than collagen. Therefore, the mass production potential and greater mechanical properties of the xetro fiber may result in these fibers becoming a new promising fiber-based material for biomedical engineering.

Multifunctional adhesive silk fibroin with blending of RGD-bioconjugated mussel adhesive protein.

Silk has recently been exploited in various fields due to its superior mechanical properties. However, this material's lack of biological functions and relatively poor biodegradation have hindered its wide use in applications related to cells and tissues. Here, we improved the overall characteristics of silkworm silk fibroin (SF) by introduction of RGD peptide-fused recombinant mussel adhesive protein (MAP-RGD). Simple blending of MAP-RGD provided not only bulk-scale adhesive ability but also microscale adhesiveness to cells and various biomolecules. MAP-RGD-blended SF fibers supported enhanced adhesion, proliferation, and spreading of mammalian cells as well as the efficient attachment of biomolecules, including carbohydrate and protein. In addition, the hydrophilicity, swelling, and biodegradability of the MAP-RGD-blended SF material were improved without notable hampering of the original mechanical properties of SF. Therefore, the adhesive silk fibrous scaffold could be successfully used in diverse biomedical engineering applications.

Diverse silk and silk-like proteins derived from terrestrial and marine organisms and their applications.

Organisms develop unique systems in a given environment. In the process of adaptation, they employ materials in a clever way, which has inspired mankind extensively. Understanding the behavior and material properties of living organisms provides a way to emulate these natural systems and engineer various materials. Silk is a material that has been with human for over 5000 years, and the success of mass production of silkworm silk has realized its applications to medical, pharmaceutical, optical, and even electronic fields. Spider silk, which was characterized later, has expanded the application sectors to textile and military materials based on its tough mechanical properties. Because silk proteins are main components of these materials and there are abundant creatures producing silks that have not been studied, the introduction of new silk proteins would be a breakthrough of engineering materials to open innovative industry fields. Therefore, in this review, we present diverse silk and silk-like proteins and how they are utilized with respect to organism's survival. Here, the range of organisms are not constrained to silkworms and spiders but expanded to other insects, and even marine creatures which produce silk-like proteins that are not observed in terrestrial silks. This viewpoint broadening of silk and silk-like proteins would suggest diverse targets of engineering to design promising silk-based materials. STATEMENT OF SIGNIFICANCE: Silk has been developed as a biomedical material due to unique mechanical and chemical properties. For decades, silks from various silkworm and spider species have been intensively studied. More recently, other silk and silk-like proteins with different sequences and structures have been reported, not only limited to terrestrial organisms (honeybee, green lacewing, caddisfly, and ant), but also from marine creatures (mussel, squid, sea anemone, and pearl oyster). Nevertheless, there has hardly been well-organized literature on silks from such organisms. Regarding the relationship among sequence-structure-properties, this review addresses how silks have been utilized with respect to organism's survival. Finally, this information aims to improve the understanding of diverse silk and silk-like proteins which can offer a significant interest to engineering fields.

Novel In Silico Analyses of Repetitive Spider Silk Sequences to Understand the Evolution and Mechanical Properties of Fibrous Protein Materials.

The mechanical properties of spider silks have diverged as spiders have diversely speciated. Because the main components of silks are proteins, it is valuable to investigate their sequences. However, silk sequences have been regarded as difficult information to analyze due to their imbalance and imperfect tandem repeats (ITR). Here, an in silico approach is applied to systemically analyze a group of silk sequences. It is found that every time new spider groups emerge, unique trimer motifs appear. These trimer motifs are used to find additional clues of evolution and to determine relationships with mechanical properties. For the first time, crucial evidence is provided that shows silk sequences coevolved with spider species and the mechanical properties of their fibers to adapt to new living environments. This novel approach can be used as a platform for analyzing other groups of ITR-harboring proteins and to obtain information for the design of tailor-made fibrous protein materials.

Natural healing-inspired collagen-targeting surgical protein glue for accelerated scarless skin regeneration.

Skin scarring after deep dermal injuries is a major clinical problem due to the current therapies limited to established scars with poor understanding of healing mechanisms. From investigation of aberrations within the extracellular matrix involved in pathophysiologic scarring, it was revealed that one of the main factors responsible for impaired healing is abnormal collagen reorganization. Here, inspired by the fundamental roles of decorin, a collagen-targeting proteoglycan, in collagen remodeling, we created a scar-preventive collagen-targeting glue consisting of a newly designed collagen-binding mussel adhesive protein and a specific glycosaminoglycan. The collagen-targeting glue specifically bound to type I collagen in a dose-dependent manner and regulated the rate and the degree of fibrillogenesis. In a rat skin excisional model, the collagen-targeting glue successfully accelerated initial wound regeneration as defined by effective reepithelialization, neovascularization, and rapid collagen synthesis. Moreover, the improved dermal collagen architecture was demonstrated by uniform size of collagen fibrils, their regular packing, and a restoration of healthy tissue component. Collectively, our natural healing-inspired collagen-targeting glue may be a promising therapeutic option for improving the healing rate with high-quality and effective scar inhibition.

Dual-Color-Emitting Carbon Nanodots for Multicolor Bioimaging and Optogenetic Control of Ion Channels.

The development of intrinsically multicolor-emitting carbon nanodots (CNDs) has been one of the great challenges for their various fields of applications. Here, the controlled electronic structure engineering of CNDs is performed to emit two distinct colors via the facile surface modification with 4-octyloxyaniline. The so-called dual-color-emitting CNDs (DC-CNDs) can be stably encapsulated within poly(styrene-co-maleic anhydride) (PSMA). The prepared water-soluble DC-CNDs@PSMA can be successfully applied to in vitro and in vivo dual-color bioimaging and optogenetics. In vivo optical imaging can visualize the biodistribution of intravenously injected DC-CNDs@PSMA. In addition, the light-triggered activation of ion channel, channelrhodopsin-2, for optogenetic applications is demonstrated. As a new type of fluorophore, DC-CNDs offer a big insight into the design of charge-transfer complexes for various optical and biomedical applications.

Upconversion Nanoparticles/Hyaluronate-Rose Bengal Conjugate Complex for Noninvasive Photochemical Tissue Bonding.

The recent progress in photonic nanomaterials has contributed greatly to the development of photomedicines. However, the finite depth of light penetration is still a serious limitation, constraining their clinical applications. Here, we developed a poly(allylamine) (PAAm)-modified upconversion nanoparticle/hyaluronate-rose bengal (UCNP/PAAm/HA-RB) conjugate complex for photochemical bonding of deep tissue with near-infrared (NIR) light illumination. Compared to the conventional invasive treatment via suturing and stapling, the UCNP/PAAm/HA-RB conjugate complex could be noninvasively delivered into the deep tissue and accelerate the tissue bonding upon NIR light illumination. HA in the outer layer of the complex facilitated the penetration of RB into the collagen layer of the dermis. The NIR light triggered UCNP of NaYF4: Yb/Er (Y:Yb:Er = 78:20:2) in the complex to illuminate visible green light under the skin tissue. The activated RB in the HA-RB conjugate by the green light induced radical formation for the cross-linking of incised collagen matrix. An in vitro light propagation test and collagen fibrillogenesis analysis, an in vivo animal tissue bonding test, and an ex vivo tensile strength test of dissected skin tissues confirmed the successful photochemical tissue bonding effect of the UCNP/PAAm/HA-RB conjugate complex.

Aquatic proteins with repetitive motifs provide insights to bioengineering of novel biomaterials.

Proteins with repetitive motifs play vital structural and adhesive functions in nature. Some repeat proteins in particular have adapted to harsh aquatic surroundings to support the survival and reproduction of organisms. Significant effort has been made to identify aquatic repeat proteins with attractive properties and functions to be used as novel biomaterials. Examples of such proteins include matrix proteins from pearl oysters, minicollagens from sea anemones, cement proteins from sandcastle worms, and byssal proteins from marine mussels. Here, several repetitive motifs from aquatic proteins are reviewed, and their characteristic properties are linked to practical uses in three aspects of aquatic life: defense, shelter, and attachment. Some repetitive motifs interact with minerals and consequently generate strong outer cover of shells, and some motifs relate with sticky nature, which contribute to organisms' habitation by adhering themselves in harsh aquatic environments. Other motifs, such as silk- or collagen-like motifs, are also involved in structural rigidity as shown in mussel's byssus and egg membrane. Thus, understanding aquatic repetitive motifs will provide clues about biomedical and biotechnological applications of engineered biomaterials in wet environments.