Tyrosine Mutation in the Characteristic Motif of the Amorphous Region of Spidroin for Self-Assembly Capability Enhancement.

Spidroin, with robust mechanical performance and good biocompatibility, could fulfill broad applications in material science and biomedical fields. Development of miniature spidroin has made abundant fiber production economically feasible, but the mechanical properties of artificial silk still fall short of natural silk. The mechanism behind mechanical properties of spidroin usually focuses on ß-microcrystalline regions; the effect of amorphous regions was barely studied. In this study, residue tyrosines (Y) were designed to replace asparagine (N)/glutamic acid (Q) in the characteristic motifs (GGX)n in amorphous regions for performance enhancement of spidroin; the mutants presented lower free energy and significantly exhibited stronger van der Waals and electrostatic interactions, which might result from π-π stacking interactions between the phenyl rings in the side chain of tyrosine. Additionally, the soluble expressions of wild-type spidroin and mutant spidroin were achieved when heterologously expressed in E. coli, with yields of 560 mg/L (2REP), 590 mg/L (2REPM), 240 mg/L (4REP), and 280 mg/L (4REPM). Significantly, secondary structure analysis confirmed that the mutant spidroin more avidly forms more ß-sheets than the wild-type spidroin, and aggregation morphology suggested that mutant spidroin displayed better self-assembly capacity and was easier to form artificial spider silk fibers; in particular, self-assembled 4REPM nanofibrils had an average modulus of 11.2 ± 0.35 GPa, about 2 times higher than self-assembled B. mori silk nanofibrils and almost the same as that of native spider dragline silk fibers (10-15 GPa). Thus, we first demonstrated a new influence mechanism of the amorphous region's characteristic motif on the self-assembly and material properties of spidroin. Our study provides a reference for the design of high-performance material proteins and their heterologous preparation.

Kirigami-inspired artificial spidroin microneedles for wound patches.

Intelligent wound management has important potential for promoting the recovery of chronic wounds caused by diabetes. Here, inspired by the field of kirigami, smart patterned high-stretch microneedle dressings (KPMDs) based on gene-modified spider silk proteins were developed to achieve sensitive biochemical and physiological sensing. The spider silk protein (spidroin) has excellent tensile properties, ductility, toughness and biocompatibility. Notably, the kirigami method-prepared kirigami structure of the spidroin MN dressing had a high tensile strength , while its ductility reached approximately 800 %. Moreover, the unique optical properties of photonic crystals allow for fluorescence enhancement, providing KPMD with color-sensitive properties suitable for wound management and clinical guidance. Furthermore, to improve the sensitivity of KPMD-s to motion monitoring, a microelectronic matrix was integrated on its surface. These distinct material properties suggest that this research lays the foundation for a new generation of high-performance biomimetic diatomaceous earth materials for application.

Efficient Biosynthetic Fabrication of Spidroins with High Spinning Performance.

The unique 3D structure of spider silk protein (spidroin) determines the excellent mechanical properties of spidroin fiber, but the difficulty of heterologous expression and poor spinning performance of recombinant spider silk protein limit its application. A high-yield low-molecular-weight biomimetic spidroin (Amy-6rep) is obtained by sequence modification, and its excellent spinning performance is verified by electrospinning it for use as a nanogenerator. Amy-6rep increases the highly fibrogenic microcrystalline region in the core repeat region of natural spidroin with limited sequence length and replaces the polyalanine sequence with an amyloid polypeptide through structural similarity. Due to sequence modification, the expression of Amy-6rep increased by ≈200%, and the self-assembly performance of Amy-6rep significantly increased. After electrospinning with Amy-6rep, the nanofibers exhibit good tribopower generation capacity. In this paper, a biomimetic spidroin sequence design with high yield and good spinning performance is reported, and a strategy for electrospinning to produce an artificial nanogenerator is explored.

[Primary Structure Characterization and Biosynthesis of Spider Silk Proteins for Multifunctional Biomaterials].

Spider silk is a natural fiber known as "biosteel" with the strongest composite performance, such as high tensile strength and toughness. It is also equipped with excellent biocompatibility and shape memory ability, thus shows great potential in many fields such as biomedicine and tissue engineering. Spider silk is composed of macromolecular spidroin with rich structural diversity. The characteristics of the primary structure of natural spidroin, such as the high repeatability of amino acids in the core repetitive region, the high content of specific amino acids, the large molecular weight, and the high GC content of the spidroin gene, have brought great difficulties in heterologous expression. This review discusses focuses on the relationship between the featured motifs of the microcrystalline region in the repetitive unit of spidroin and its structure, as well as the spinning performance and the heterologous expression. The optimization design for the sequence of spidroin combined with heterologous expression strategy has greatly promoted the development of the biosynthesis of spider silk proteins. This review may facilitate the rational design and efficient synthesis of recombinant spidroin.

Patterned Duplex Fabric Based on Genetically Modified Spidroin for Smart Wound Management.

The treatment of diabetic wounds remains a great challenge for the medical community. Here, a smart patterned DNA double helix (duplex)-like fabric based on genetically modified spider silk protein (PDF-S) which is inspired by soft plant tendrils, is proposed for diabetic wound treatment. Benefiting from spider silk protein (spidroin); PDF-S is equipped with high strength; high toughness, and excellent biocompatibility. Notably, the fabric crimped through the biomimetic DNA double-helix-like structure can effectively adapt to tensile impact and the maximum stretch rate reaches 1500%. A pattern-based microfluidic channel of PDF-S allowed wound secretion to flow spontaneously through the channel. Meanwhile; due to the optical properties of the introduced photonic crystal structure; PDF-S is equipped with fluorescence enhancement properties; enabling PDF-S to display color-sensitive behavior suitable for wound monitoring and guiding clinical treatment. In addition, to enable sensitive motion monitoring, microelectronic circuits are integrated on the surface of the PDF-S. These unique material features suggest that this study will lead to a new generation of biomimetic artificial spider silk materials for design and application in the biomedical field.