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We showed previously that a rigid and fibrous-structured cationic coiled-coil artificial protein had cell-penetrating activity that was significantly greater when compared with a less-structured cell-penetrating peptide. Nanomaterials with anisotropic structures often show aspect-ratio-dependent unique physicochemical properties, as well as cell-penetrating activities. In this report, we have designed and demonstrated the cell-penetrating activity of a shorter cationic coiled-coil protein. An aspect ratio at 4.5:1 was found to be critical for ensuring that the cationic coiled-coil protein showed strong cell-penetrating activity. At an aspect ratio of 3.5:1, the cationic coiled-coil protein showed cell-penetrating activity that was similar to a less-structured short cationic cell-penetrating peptide. Interestingly, at an aspect ratio of 4:1, the cationic coiled-coil protein exhibited intermediate cell-penetrating activity. These findings should aid in the principle design of intracellular drug delivery carriers including coiled-coil artificial proteins, their derivatives, and α-helical cell-penetrating peptides as well as provide a framework for developing synthetic nanomaterials, such as metal nanorods and synthetic polymers.

Numerous cationic peptides that penetrate cells have been studied intensively as drug delivery system carriers for cellular delivery. However, cationic molecules tend to be cytotoxic and cause inflammation, and their stability in the blood is usually low. We have previously demonstrated that a rigid and fibrous cationic coiled-coil protein exhibited cell-penetrating ability superior to that of previously reported cell-penetrating peptides. Making use of structural properties, here we describe the cell-penetrating activity of a rigid and fibrous coiled-coil protein with a noncationic surface. A fibrous coiled-coil protein of pI 6.5 penetrated 100% of the cells tested in vitro at a concentration of 500 nM, which is comparable to that of previously reported cell-penetrating peptides. We also investigated the effect of cell-strain dependency and short-term cytotoxicity.

Molecules with structural anisotropy and rigidity, such as asbestos, demonstrate high cell-penetrating activity but also high toxicity. Here we synthesize a biodegradable, rigid, and fibrous artificial protein, CCPC 140, as a potential vehicle for cellular delivery. CCPC 140 penetrated 100% of cells tested in vitro, even at a concentration of 3.1 nM-superior to previously reported cell-penetrating peptides. The effects of cell-strain-dependency and aspect ratio on the cell-penetrating activity of CCPC 140 were also investigated.

The first six peptides of multifunctional titanium binding peptide-1 bestowed recombinant L-ferritin, minT1-LF, was genetically engineered and used to fabricate multilayered nanoparticle architecture. The multifunctionality of minT1-LF enables specific binding of nanoparticle-accommodated minT1-LF to the silicon substrate surface and wet biochemical fabrication of gate oxide layer by its biomineralization activity. Three-dimensional (3D) nanoparticle architecture with multilayered structure was fabricated by the biological layer-by-layer method and embedded in a metal oxide-semiconductor device structure as a charge storage node of a flash memory device. The 3D-integrated multilayered nanoparticle architecture successfully worked as a charge storage node in flash memory devices that exhibited improved charge storage capacity compared with that of a conventional monolayer structure device.

We present a novel method for preparing a silica carrier for the sustained release of a proteinaceous pharmaceutical. This method makes use of the silicification activity of the protein itself, which autonomously formed a protein-silica composite upon simple incubation with a silica precursor. The composite was dissolved, and the encapsulated protein was released into a culture medium, thereby sustaining the protein's activity for a long period of time.

The interactions of ferritins fused with a Ti-recognizing peptide (RKLPDA) and their mutants with titanium oxide substrates were explored with an atomic force microscope (AFM). The amino acid sequence of the peptide was systematically modified to elucidate the role of each amino acid residue in the specific interaction. Force measurements revealed a clear correlation among the sequences in the N-terminal domain of ferritin, surface potentials, and long-range electrostatic interactions. Measurements of adhesion forces clearly revealed that hydrogen bonds take part in the specific binding as well as the electrostatic interaction between charged residues and surface charges of Ti oxides. Moreover, our results indicated that not only the charged and polar residues but also a neutral residue (proline) govern the strength of the specific binding, with the order of the residues also being significant. These results demonstrate that the local structure of the peptide governs the special arrangement of charged residues and strongly affects the strength of the bindings.

A two-dimensional hexagonally close-packed (2D-HCP) array of ferritin molecules with a nanoparticle core was fabricated directly on a carbonaceous solid substrate by genetically modifying the outer surface of the ferritin with carbonaceous materials-specific binding peptides. The displayed peptides endowed ferritins with a specific protein-substrate interaction and masked the strong nonspecific interaction. The specific interaction was weak enough to allow ferritins to be rearranged as well as an attractive protein-protein interaction that could be adjusted by selecting the buffer conditions. This method not only produced 2D-HCP arrays of ferritin but also 2D-ordered arrays of independent inorganic nanoparticles after protein elimination that can be applied to floating gate memories.

Numerous peptide aptamers that recognize inorganic materials have been isolated using in vitro peptide evolution systems. However, it remains unknown how peptides interact with inorganic materials or how specific those interactions are. We, therefore, assessed the target specificities of the peptide aptamer TBP-1 (RKLPDAPGMHTW) by monitoring its ability to bind 10 different metals. We found that phages displaying TBP-1 bound to Ti, Si, and Ag surfaces but not to Au, Cr, Pt, Sn, Zn, Cu, or Fe. As previously seen with Ti, binding to Si and Ag was diminished by R1A, P4A, or D5A mutation, suggesting that the same molecular mechanism underlies TBP-1 binding to all three materials. We also observed that a synthetic TBP-1 peptide mediated mineralization of both silica and Ag. It, thus, appears that although the overall chemical characteristics of Ti, Si, and Ag surfaces are dissimilar, they share a common subnanometric structure that is recognized by TBP-1.