Short synthetic peptide amphiphiles have recently been explored as effective nanobiomaterials in applications ranging from controlled gene and drug release, skin care, nanofabrication, biomineralization, membrane protein stabilization to 3D cell culture and tissue engineering. This range of applications is heavily linked to their unique nanostructures, remarkable simplicity and biocompatibility. Some peptide amphiphiles also possess antimicrobial activities whilst remaining benign to mammalian cells. These attractive features are inherently related to their selective affinity to different membrane interfaces, high capacity for interfacial adsorption, nanostructuring and spontaneous formation of nano-assemblies. Apart from sizes, the primary sequences of short peptides are very diverse as they can be either biomimetic or de novo designed. Thus, their self-assembling mechanistic processes and the nanostructures also vary enormously. This critical review highlights recent advances in studying peptide amphiphiles, focusing on the formation of different nanostructures and their applications in diverse fields. Many interesting features learned from peptide self-organisation and hierarchical templating will serve as useful guidance for functional materials design and nanobiotechnology (123 references).

Amphiphilic peptides A(3)K, A(6)K, and A(9)K displayed an increasing propensity for nanoaggregation with increasing the size of hydrophobic alanine moiety, and the size and shape of the aggregates showed a steady transition from loose peptide stacks formed by A(3)K, long nanofibers by A(6)K, to short and narrow nanorods by A(9)K. This size and shape transition was broadly consistent with the trend predicted from interfacial packing and curvature change if these peptide surfactants were treated as conventional surfactants. The antibacterial capacity, defined by the killing of percentage of bacteria in a given time and peptide concentration, showed a strong correlation to peptide hydrophobicity, evident from both microscopic and fluorescence imaging studies. For A(9)K, the power for membrane permeation and bacterial clustering intensified with peptide concentration and incubation time. These results thus depict a positive correlation between the propensity for self-assembly of the peptides, their membrane penetration power, and bactericidal capacity. Although the exposure of A(9)K to a preformed DPPC membrane bilayer showed little structural disturbance, the same treatment to the preformed DPPG membrane bilayer led to substantial disruption of model membrane structure, a trend entirely consistent with the high selectivity observed from membrane hemolytic studies.

Peptide amphiphiles readily self-assemble into a variety of nanostructures, but how molecular architectures affect the size and shape of the nanoaggregates formed is not well understood. From a combined TEM and AFM study of a series of cationic peptide surfactants AmK (m = 3, 6, and 9), we show that structural transitions (sheets, fibers/ worm-like micelles, and short rods) can be induced by increasing the length of the hydrophobic peptide region. The trend can be interpreted using the molecular packing theory developed to describe surfactant structural transitions, but the entropic gain, decreased CAC, and increased electrostatic interaction associated with increasing the peptide hydrophobic chain need to be taken into account appropriately. Our analysis indicates that the trend in structural transitions observed from AmK peptide surfactants is opposite to that obtained from conventional monovalent ionic surfactants. The outcome reflects the dominant role of hydrophobic interaction between the side chains opposed by backbone hydrogen bonding and electrostatic repulsion between lysine side chains.

Many de novo designed amphiphilic peptides capable of self-assembly and further structural templating into hierarchical organizations such as nanofibers and gels carry more than 10 amino acid residues. A curious question is now raised about the minimal size that is required for initiating amphiphilically driven nanostructuring. In this work, we show that ultrashort peptides I 3 K and L 3 K could readily self-assemble into stable nanostructures. While L 3 K formed spherical nanospheres with diameters of ∼10-15 nm, I 3 K self-assembled into nanotubes with diameters of ∼10 nm and lengths of >5 μm. I 3 K nanotubes were very smooth and carried defined pitches of twisting. The difference could arise from the different β-sheet promoting power between isoleucine and leucine, suggesting that while hydrophobic interaction was dominant in the formation of L 3 K nanospheres hydrogen bonding governed the templating of antiparallel β-sheets and the subsequent formation of I 3 K nanotubes. Because of their extreme stability against heating or exposure to organic solvents, I 3 K nanotubes were used as templates for silicification from the hydrolysis of organosilicate precursors using TEOS (tetraethoxysilane). The lysine groups on the inner and outer nanotube surfaces worked to catalyze silicification, leading to the formation of silica nanotubes, which is evident from both AFM and TEM imaging. The formation of interesting nanotubes and nanospheres as demonstrated from very short peptide amphiphiles is significant for further exploration of their use in technological applications.

Peptide self-assembly is of direct relevance to protein science and bionanotechnology, but the underlying mechanism is still poorly understood. Here, we demonstrate the distinct roles of the noncovalent interactions and their impact on nanostructural templating using carefully designed hexapeptides, I2K2I2, I4K2, and KI4K. These simple variations in sequence led to drastic changes in final self-assembled structures. β-sheet hydrogen bonding was found to favor the formation of one-dimensional nanostructures, such as nanofibrils from I4K2 and nanotubes from KI4K, but the lack of evident β-sheet hydrogen bonding in the case of I2K2I2 led to no nanostructure formed. The lateral stacking and twisting of the β-sheets were well-linked to the hydrophobic and electrostatic interactions between amino acid side chains and their interplay. For I4K2, the electrostatic repulsion acted to reduce the hydrophobic attraction between β-sheets, leading to their limited lateral stacking and more twisting, and final fibrillar structures; in contrast, the repulsive force had little influence in the case of KI4K, resulting in wide ribbons that eventually developed into nanotubes. The fibrillar and tubular features were demonstrated by a combination of cryogenic transmission electron microscopy (cryo-TEM), negative-stain transmission electron microscopy (TEM), and small-angle neutron scattering (SANS). SANS also provided structural information at shorter scale lengths. All atom molecular dynamics (MD) simulations were used to suggest possible molecular arrangements within the β-sheets at the very early stage of self-assembly.

Interfacial adsorption of a mouse monoclonal antibody (type IgG1, anti-beta-hCG) at the hydrophilic silicon oxide/water interface has been studied by spectroscopic ellipsometry and neutron reflection, followed by assessment of binding of a hormonal antigen, human chorionic gonadotrophin (hCG), onto the adsorbed antibody molecules. The amount of adsorption reached a maximum around the isoelectric pH (IP) of 6 for the antibody; this pH-dependent pattern could be altered by increasing salt concentration, a trend also observed for other proteins. Neutron reflection revealed the formation of a 40 A uniform layer from the adsorbed antibody, indicating a flat-on orientation. The subsequent hCG binding showed that the molar ratio of hCG bound to antibody at the interface was as high as 0.7 at low surface coverage of antibody and decreased with increasing surface antibody concentration. The results point to an increasing extent of steric hindrance to hCG access with increasing packing density of antibody molecules on the surface. Comparison with previously published crystal structure studies suggests twisting of the variable region to allow access of the antigen. The binding of hCG was also found to be pH-dependent with its maximum around the IP, if the ionic strength of the solution was low (20 mM). However, if the ionic strength was increased to 200 mM, then hCG binding was influenced by a combination of steric hindrance and electrostatic interaction between the antigen and the surface. These results are highly relevant to the improvement of the performance of biotechnologies such as fertility test pads and biosensors based on antibody immobilization.

The interplay between hydrogen bonding, hydrophobic interaction and the molecular geometry of amino acid side-chains is crucial to the development of nanostructures of short peptide amphiphiles. An important step towards developing their practical use is to understand how different amino acid side-chains tune hydrophobic interaction and hydrogen bonding and how this process leads to the control of the size and shape of the nanostructures. In this study, we have designed and synthesized three sets of short amphiphilic peptides (I(3)K, LI(2)K and L(3)K; L(3)K, L(4)K and L(5)K; I(3)K, I(4)K and I(5)K) and investigated how I and L affected their self-assembly in aqueous solution. The results have demonstrated a strong tendency of I groups to promote the growth of β-sheet hydrogen bonding and the subsequent formation of nanofibrillar shapes. All I(m)K (m = 3-5) peptides assembled into nanofibers with consistent β-sheet conformation, whereas the nanofiber diameters decreased as m increased due to geometrical constraint in peptide chain packing. In contrast, L groups had a weak tendency to promote β-sheet structuring and their hydrophobicity became dominant and resulted in globular micelles in L(3)K assembly. However, increase in the number of hydrophobic sequences to L(5)K induced β-sheet conformation due to the cooperative hydrophobic effect and the consequent formation of long nanofibers. The assembly of L(4)K was, therefore, intermediate between L(3)K and L(5)K, similar to the case of LI(2)K within the set of L(3)K, LI(2)K and I(3)K, with a steady transition from the dominance of hydrophobic interaction to hydrogen bonding. Thus, changes in hydrophobic length and swapping of L and I can alter the size and shape of the self-assembled nanostructures from these simple peptide amphiphiles.

We report a new class of cationic amphiphilic peptides with short sequences, G(IIKK)(n)I-NH(2) (n = 1-4), that can kill Gram-positive and Gram-negative bacteria as effectively as several well-known antimicrobial peptides and antibiotics. In addition, some of these peptides possess potent antitumor activities against cancer cell lines. Moreover, their hemolytic activities against human red blood cells (hRBCs) remain remarkably low even at some 10-fold bactericidal minimum inhibitory concentrations (MICs). When bacteria or tumor cells are cocultured with NIH 3T3 fibroblast cells, G(IIKK)(3)I-NH(2) showed fast and strong selectivity against microbial or tumor cells, without any adverse effect on NIH 3T3 cells. The high selectivity and associated features are attributed to two design tactics: the use of Ile residues rather than Leu and the perturbation of the hydrophobic face of the helical structure with the insertion of a positively charged Lys residue. This class of simple peptides hence offers new opportunities in the development of cost-effective and highly selective antimicrobial and antitumor peptide-based treatments.