Self‐assembling peptides that are capable of adopting β‐sheet structures can generate nanofibers that lead to hydrogel formation. Herein, to tune the supramolecular morphologies, mechanical properties, and stimuli responses of the hydrogels, we investigated glycine substitution in a β‐sheet‐forming amphiphilic peptide. Glycine substitution generally enhances conformational flexibility. Indeed, glycine substitution in an amphiphilic peptide weakened the hydrogels or even inhibited the gelation. However, unexpectedly, glycine substitution at the center of the peptide molecule significantly enhanced the hydrogel stiffness. The central glycine substitution affected the molecular packing and led to twisted β‐sheet structures and to nanofiber bundling, which likely led to the stiffened hydrogel. Importantly, the supramolecular structures were accurately predicted by molecular dynamics simulations, demonstrating the helpfulness of these techniques for the identification of self‐assembling peptides. The hydrogel formed by the amphiphilic peptide with the central glycine substitution had cell adhesive function, and showed a reversible thermal gel‐to‐sol transition. Thus, glycine substitution is effective in modulating self‐assembling structures, rheological properties, and dynamics of biofunctional self‐assembling peptides.

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Cell adhesion is a fundamental biological process involved in a wide range of cellular and biological activity. Integrin−ligand binding is largely responsible for cell adhesion with an extracellular matrix, and the RGD sequence is an epitope in ligand proteins such as fibronectin. The extracellular matrix consists of fibrous proteins with embedded ligands for integrins. Such a biological architecture has been reconstructed for biochemical, pharmaceutical, and biomaterial studies using artificial supramolecular systems to reproduce cell adhesion functionality, and fiber-forming self-assembling peptides containing RGD are one such promising material for this purpose. In this study, using RADA16 as a model fiber-forming peptide, a series of RGD-containing variants have been synthesized by the replacement of one alanine with glycine at different positions, in which all the variants consist of identical amino acid components. The position of the RGD unit influenced the supramolecular self-assembly of the amphiphilic peptide to inhibit β-sheet formation (A6G) or twist the molecular alignment in β-sheet-type assemblies (A10G and A14G). Furthermore, A10G and A14G formed assembled nanofibers, which afforded hydrogels with higher viscoelasticities than other RGD-containing variants. In contrast to A10G and A14G, which exhibit substantial cell adhesion functionality, the cell adhesion efficiencies of the other RGD-containing variants were significantly reduced. This suggests that the higher order structure could strongly influence the cell adhesion functionality of RGD-containing supramolecular nanofibers.

Invited for the cover of this issue is the group of Takahiro Muraoka at Tokyo University of Agriculture and Technology and collaborators. The image depicts bundled nanofiber formation by self‐assembly of a β‐sheet‐forming an amphiphilic peptide with a central glycine substitution. Read the full text of the article at 10.1002/chem.201902083.

For the development of antiviral agents to eliminate hepatitis B virus (HBV), it is essential to establish an HBV cell culture system that can easily monitor HBV infection. Here, we created a novel HBV infection monitoring system using a luminescent 11‐amino acid reporter, the high‐affinity subunit of nano‐luciferase binary technology (HiBiT). The HiBiT‐coding sequence was inserted at the N‐terminus of preS1 in a 1.2‐fold plasmid encoding a genotype C HBV genome. After transfection of HepG2 cells with this HiBiT‐containing plasmid, the supernatant was used to prepare a recombinant cell culture‐derived virus (HiBiT‐HBVcc). Primary human hepatocytes (PXB) were inoculated with HiBiT‐HBVcc. Following inoculation, intracellular and extracellular HiBiT activity and the levels of various HBV markers were determined. Reinfection of naive PXB cells with HiBiT‐HBVcc prepared from HiBiT‐HBVcc‐infected PXB cells was analyzed. When PXB cells were infected with HiBiT‐HBVcc at several titers, extracellular HiBiT activity was detected in a viral titer‐dependent manner and was correlated with intracellular HiBiT activity. Inhibitors of HBV entry or replication suppressed extracellular HiBiT activity. Viral DNA, RNA, and proteins were detectable, including covalently closed circular DNA, by Southern blot analysis. The synthesis of relaxed‐circular DNA from single‐stranded DNA in HiBiT‐HBV decreased to one third of that of wild‐type HBV, and the infectivity of HiBiT‐HBVcc decreased to one tenth of that of wild‐type HBVcc. HiBiT‐HBVcc prepared from PXB cells harboring HiBiT‐HBV was able to infect naive PXB cells. Conclusions : Recombinant HiBiT‐HBV can undergo the entire viral life cycle, thus facilitating high‐throughput screening for HBV infection in vitro using supernatants. This system will be a powerful tool for developing antiviral agents.