A series of nine, frustrated, multidomain peptides is described in which forces favoring self-assembly into a nanofiber versus those favoring disassembly could be easily modified. The peptides are organized into an ABA block motif in which the central B block is composed of alternating hydrophilic and hydrophobic amino acids (glutamine and leucine, respectively). This alternation allows the amino acid side chains to segregate on opposite sides of the peptide backbone when it is in a fully extended beta-sheet conformation. In water, packing between two such peptides stabilizes the extended conformation by satisfying the desire of the leucine side chains to exclude themselves from the aqueous environment. Once in this conformation intermolecular backbone hydrogen bonding can readily take place between additional peptides eventually growing into high aspect ratio fibers. B block assembly may continue infinitely or until monomeric peptides are depleted from solution which results in an insoluble precipitate. Block A consists of a variable number of positively charged lysine residues whose electrostatic repulsion at pH 7 works against the desire of the B block to assemble. Here we show that balancing the forces of block A against B allows the formation of controlled length, individually dispersed, and fully soluble nanofibers with a width of 6 +/- 1 nm and length of 120 +/- 30 nm. Analysis by infrared, circular dichroism, and vitreous ice cryo-transmission electron microscopy reveals that the relative sizes of blocks A and B dictate the peptide secondary structure which in turn controls the resulting nanostructure. The system described epitomizes the use of molecular frustration in the design of finite self-assembled structures. These materials, and ones based on their architecture, may find applications where nanostructured control over fiber architecture and chemical functionality is required.

An important goal in supramolecular chemistry is to achieve controlled self-assembly of molecules into well-defined nanostructures and the subsequent control over macroscopic properties resulting from the formation of a nanostructured material. Particularly important to our lab is control over viscoelasticity and bioactivity. Recently we described a multidomain peptide motif that can self-assemble into nanofibers 2 x 6 x 120 nm. In this work we describe how sequence variations in this general motif can be used to create nanofibrous gels that have storage moduli, which range over 2 orders of magnitude and can undergo shear thinning and shear recovery while at the modest concentration of 1% by weight. Gel formation is controlled by addition of oppositely charged multivalent ions such as phosphate and magnesium and can be carried out at physiological pH. We also demonstrate how maximum strength can be obtained via covalent capture of the nanofibers through disulfide bond formation. Together these hydrogel properties are ideally suited as injectable materials for drug and cell delivery.

The R-helical coiled coil is one of the best-studied and most well-understood protein folding motifs. In particular, the coiled coil can be made to self-assemble into a nanofibrous architecture with many potential applications in biomimetic engineering and elsewhere. The key to the assembly of such nanofibers has been the formation of "sticky ended" dimers through careful selection of electrostatically charged amino acids. In this work, we demonstrate for the first time that sticky ended dimers are not a prerequisite for R-helical coiled coil nanofiber formation. In contrast, we show that blunt-ended dimers are able to form nanofibers with a uniform diameter of 4 nm while being hundreds of nanometers in length. Furthermore, the length and lateral packing can be controlled through selection of amino acids not involved in the coiled coil interface.

Generating stable, multi-functional organic nanocarriers will have a significant impact on drug formulation. However, it remains a significant challenge to generate organic nanocarriers with a long circulation half-life, effective tumor penetration and efficient clearance of metabolites. We have advanced this goal by designing a new family of amphiphiles based on coiled-coil 3-helix bundle forming peptide-poly(ethylene glycol) conjugates. The amphiphiles self-assemble into monodisperse micellar nanoparticles, 15 nm in diameter. Using the 3-helix micelles, a drug loading of ~8 wt% was obtained using doxorubicin (DOX) and the micelles showed minimal cargo leakage after 12 hours of incubation with serum proteins at 37°C. In vivo pharmacokinetics studies using positron emission tomography (PET) showed a circulation half-life of 29.5 hrs and minimal accumulation in the liver and spleen. The demonstrated strategy, by incorporating unique protein tertiary structure in the headgroup of an amphiphile, opens new avenues to generate organic nanoparticles with tunable stability, ligand clustering and controlled disassembly to meet current demands in nanomedicine.

Dental caries remains one of the most prevalent infectious diseases in the world. So far, available treatment methods rely on the replacement of decayed soft and mineralized tissue with inert biomaterials alone. As an approach to develop novel regenerative strategies and engineer dental tissues, two dental stem cell lines were combined with peptide-amphiphile (PA) hydrogel scaffolds. PAs self-assemble into three-dimensional networks of nanofibers, and living cells can be encapsulated. Cell-matrix interactions were tailored by incorporation of the cell adhesion sequence RGD and an enzyme-cleavable site. SHED (stem cells from human exfoliated deciduous teeth) and DPSC (dental pulp stem cells) were cultured in PA hydrogels for 4 weeks using different osteogenic supplements. Both cell lines proliferate and differentiate within the hydrogels. Histologic analysis shows degradation of the gels and extracellular matrix production. However, distinct differences between the two cell lines can be observed. SHED show a spindle-shaped morphology, high proliferation rates, and collagen production, resulting in soft tissue formation. In contrast, DPSC reduce proliferation, but exhibit an osteoblast-like phenotype, express osteoblast marker genes, and deposit mineral. Since the hydrogels are easy to handle and can be introduced into small defects, this novel system might be suitable for engineering both soft and mineralized matrices for dental tissue regeneration.

Despite increasing demands to employ amphiphilic micelles as nanocarriers and nanoreactors, it remains a significant challenge to simultaneously reduce the particle size and enhance the particle stability. Complementary to covalent chemical bonding and attractive intermolecular interactions, entropic repulsion can be incorporated by rational design in the headgroup of an amphiphile to generate small micelles with enhanced stability. A new family of amphiphilic peptide-polymer conjugates is presented where the hydrophilic headgroup is composed of a 3-helix coiled-coil with poly(ethylene glycol) attached to the exterior of the helix bundle. When micelles form, the PEG chains are confined in close proximity and are compressed to act as a spring to general lateral pressure. The formation of 3-helix bundles determines the location and the directionalities of the force vector of each PEG elastic spring so as to slow down amphiphile desorption. Since each component of the amphiphile can be readily tailored, these micelles provide numerous opportunities to meet current demands for organic nanocarriers with tunable stability in life science and energy science. Furthermore, present studies open new avenues to use energy arising from entropic polymer chain deformation to self-assemble energetically stable single nanoscopic objects, much like repulsion that stabilizes bulk assemblies of colloidal particles.

This work demonstrates a design strategy to optimize antimicrobial peptides with an ideal balance of minimal cytotoxicity, enhanced stability, potent cell penetration and effective antimicrobial activity, which hold great promise for the treatment of intracellular microbial infections and potentially systemic anti-infective therapy.

Cellular membrane disruption induced by ␤-amyloid (A␤) peptides has been considered one of the major pathological mechanisms for Alzheimer disease. Mechanistic studies of the membrane disruption process at a high-resolution level, on the other hand, are hindered by the co-existence of multiple possible pathways, even in simplified model systems such as the phospholipid liposome. Therefore, separation of these pathways is crucial to achieve an in-depth understanding of the A␤-induced membrane disruption process. This study, which utilized a combination of multiple biophysical techniques, shows that the peptide-to-lipid (P:L) molar ratio is an important factor that regulates the selection of dominant membrane disruption pathways in the presence of 40-residue A␤ peptides in liposomes. Three distinct pathways (fibrillation with membrane content leakage, vesicle fusion, and lipid uptake through a temporarily stable ionic channel) become dominant in model liposome systems under specific conditions. These individual systems are characterized by both the initial states of A␤ peptides and the P:L molar ratio. Our results demonstrated the possibility to generate simplified A␤-membrane model systems with a homogeneous membrane disruption pathway, which will benefit high-resolution mechanistic studies in the future. Fundamentally, the possibility of pathway selection controlled by P:L suggests that the driving forces for A␤ aggregation and A␤-membrane interactions may be similar at the molecular level.