Chengkang Tang

Temperature and pH effects on biophysical and morphological properties of self-assembling peptide RADA16-I.

It has been found that the self‐assembling peptide RADA 16‐I forms a β‐sheet structure and self‐assembles into nanofibers and scaffolds in favor of cell growth, hemostasis and tissue‐injury repair. But its biophysical and morphological properties, especially for its β‐sheet and self‐assembling properties in heat‐ and pH‐denatured conditions, remain largely unclear. In order to better understand and design nanobiomaterials, we studied the self‐assembly behaviors of RADA16‐I using CD and atomic force microscopy (AFM) measurements in various pH and heat‐denatured conditions. Here, we report that the peptide, when exposed to pH 1.0 and 4.0, was still able to assume a typical β‐sheet structure and self‐assemble into long nanofiber, although its β‐sheet content was dramatically decreased by 10% in a pH 1.0 solution. However, the peptide, when exposed to pH 13.0, drastically lost its β‐sheet structure and assembled into different small‐sized globular aggregates. Similarly, the peptide, when heat‐denatured from 25 to 70 °C, was still able to assume a typical β‐sheet structure with 46% content, but self‐assembled into small‐sized globular aggregates at much higher temperature. Titration experiments showed that the peptide RADA16‐I exists in three types of ionic species: acidic (fully protonated peptide), zwitterionic (electrically neutral peptide carrying partial positive and negative charges) and basic (fully deprotonated peptide) species, called ‘super ions’. The unordered structure and β‐turn of these ‘super ions’ via hydrogen or ionic bonds, and heat Brownian motion under the above denatured conditions would directly affect the stability of the β‐sheet and nanofibers. These results help us in the design of future nanobiomaterials, such as biosensors, based on β‐sheets and environmental changes. These results also help understand the pathogenesis of the β‐sheet‐mediated neuronal diseases such as Alzheimers disease and the mechanism of hemostasis. Copyright

De Novo Design of a Bolaamphiphilic Peptide with Only Natural Amino Acids

A new self-assembling bolaamphiphilic peptide has been designed and synthesized using only natural amino acids. This simple peptide is composed of two lysines connected by 4-8 alanines to maintain the characteristics of the traditional bolaamphiphiles. Based on an irregular secondary structure, it can self-assemble into nanospheres, nanorods, or nanofibers with lengths up to micrometers. The long nanofibers can be broken into smaller fragments by sonication, however, they could reassemble into nanofibers after incubation. Furthermore, the nanostructures were shown to have considerable thermostability. This new bolaamphiphilic peptide differs from any other self-assembling peptides or bolaamphiphiles, and possibly provides a new approach to fabricate nanomaterials.

The effect of self-assembling peptide RADA16-I on the growth of human leukemia cells in vitro and in nude mice.

Nanofiber scaffolds formed by self-assembling peptide RADA16-I have been used for the study of cell proliferation to mimic an extracellular matrix. In this study, we investigated the effect of RADA16-I on the growth of human leukemia cells in vitro and in nude mice. Self-assembly assessment showed that RADA16-I molecules have excellent self-assembling ability to form stable nanofibers. MTT assay displayed that RADA16-I has no cytotoxicity for leukemia cells and human umbilical vein endothelial cells (HUVECs) in vitro. However, RADA16-I inhibited the growth of K562 tumors in nude mice. Furthermore, we found RADA16-I inhibited vascular tube-formation by HUVECs in vitro. Our data suggested that nanofiber scaffolds formed by RADA16-I could change tumor microenvironments, and inhibit the growth of tumors. The study helps to encourage further design of self-assembling systems for cancer therapy.

Molecular design and applications of self-assembling surfactant-like peptides

Self-assembling surfactant-like peptides have been explored as emerging nanobiomaterials in recent years. These peptides are usually amphiphilic, typically possessing a hydrophobicmoiety and a hydrophilicmoiety. The structural characteristics can promote many peptide molecules to self-assemble into various nanostructures. Furthermore, properties of peptide molecules such as charge distribution and geometrical shape could also alter the formation of the self-assembling nanostructures. Based on their diverse self-assembling behaviours and nanostructures, self-assembling surfactant-like peptides exhibit great potentials in many fields, including membrane protein stabilization, drug delivery, and tissue engineering. This review mainly focuses on recent advances in studying self-assembling surfactant-like peptides, introducing their designs and the potential applications in nanobiotechnology.

Self-assembling surfactant-like peptide A6K as potential delivery system for hydrophobic drugs.

Background Finding a suitable delivery system to improve the water solubility of hydrophobic drugs is a critical challenge in the development of effective formulations. In this study, we used A6K, a self-assembling surfactant-like peptide, as a carrier to encapsulate and deliver hydrophobic pyrene. Methods Pyrene was mixed with A6K by magnetic stirring to form a suspension. Confocal laser scanning microscopy, transmission electron microscopy, dynamic light scattering, atomic force microscopy, fluorescence, and cell uptake measurements were carried out to study the features and stability of the nanostructures, the state and content of pyrene, as well as the pyrene release profile. Results The suspension formed contained pyrene monomers trapped in the hydrophobic cores of the micellar nanofibers formed by A6K, as well as nanosized pyrene crystals wrapped up and stabilized by the nanofibers. The two different encapsulation methods greatly increased the concentration of pyrene in the suspension, and formation of pyrene crystals wrapped up by A6K nanofibers might be the major contributor to this effect. Furthermore, the suspension system could readily release and transfer pyrene into living cells. Conclusion A6K could be further exploited as a promising delivery system for hydrophobic drugs.

FORMATION OF REVERSED MICELLE NANORING BY A DESIGNED SURFACTANT-LIKE PEPTIDE

Designing self-assembling peptides as nanomaterials has been an attractive strategy in recent years, however, these peptides were usually studied in aqueous solutions for their self-assembling behaviors and applications. In this study, we have designed a surfactant-like peptide AGD with a wedge-like shape and studied its self-assembling behaviors in aqueous solution or nonpolar system. By analyzing the intermolecular hydrogen bond using FT-IR and characterizing the nanostructures with DLS, AFM and TEM, it was confirmed that AGD could not undergo self-assembly in aqueous solution while could self-assemble into well-ordered nanorings in nonpolar system. A molecular model has been proposed to explain how the nanorings were formed in the manner of reversed micelle. These results suggested a novel strategy to fabricate self-assembling peptide nanomaterials in nonpolar system, which could have potential applications in many fields.

Ethanol induced the formation of β-sheet and amyloid-like fibrils by surfactant-like peptide A6K.

Self‐assembly of natural or designed peptides into fibrillar structures based on β‐sheet conformation is a ubiquitous and important phenomenon. Recently, organic solvents have been reported to play inductive roles in the process of conformational change and fibrillization of some proteins and peptides. In this study, we report the change of secondary structure and self‐assembling behavior of the surfactant‐like peptide A6K at different ethanol concentrations in water. Circular dichroism indicated that ethanol could induce a gradual conformational change of A6K from unordered secondary structure to β‐sheet depending upon the ethanol concentration. Dynamic light scattering and atomic force microscopy revealed that with an increase of ethanol concentration the nanostructure formed by A6K was transformed from nanosphere/string‐of‐beads to long and smooth fibrils. Furthermore, Congo red staining/binding and thioflavin‐T binding experiments showed that with increased ethanol concentration, the fibrils formed by A6K exhibited stronger amyloid fibril features. These results reveal the ability of ethanol to promote β‐sheet conformation and fibrillization of the surfactant‐like peptide, a fact that may be useful for both designing self‐assembling peptide nanomaterials and clarifying the molecular mechanism behind the formation of amyloid fibrils. Copyright

Amphiphilic peptides as novel nanomaterials: design, self-assembly and application

Designer self-assembling peptides are a category of emerging nanobiomaterials which have been widely investigated in the past decades. In this field, amphiphilic peptides have received special attention for their simplicity in design and versatility in application. This review focuses on recent progress in designer amphiphilic peptides, trying to give a comprehensive overview about this special type of self-assembling peptides. By exploring published studies on several typical types of amphiphilic peptides in recent years, herein we discuss in detail the basic design, self-assembling behaviors and the mechanism of amphiphilic peptides, as well as how their nanostructures are affected by the peptide characteristics or environmental parameters. The applications of these peptides as potential nanomaterials for nanomedicine and nanotechnology are also summarized.

Amyloid-like staining property of RADA16-I nanofibers and its potential application in detecting and imaging the nanomaterial

Background Designer self-assembling peptide nanofibers (SAPNFs) as a novel kind of emerging nanomaterial have received more and more attention in the field of nanomedicine in recent years. However, a simple method to monitor and image SAPNFs is still currently absent. Methods RADA16-I, a well-studied ionic complementary peptide was used as a model to check potential amyloid-like staining properties of SAPNFs. Thioflavin-T (ThT) and Congo red (CR) as specific dyes for amyloid-like fibrils were used to stain RADA16-I nanofibers in solution, combined with drugs or cells, or injected in vivo as hydrogels. Fluorescent spectrometry and fluorescent microscopy were used to check ThT-binding property, and polarized light microscopy was used to check CR-staining property. Results ThT binding with the nanofibers showed enhanced and blue-shifted fluorescence, and specific apple-green birefringence could be observed after the nanofibers were stained with CR. Based on these properties we further showed that ThT-binding fluorescence intensity could be used to monitor the forming and changing of nanofibers in solution, while fluorescent microscopy and polarized light microscopy could be used to image the nanofibers as material for drug delivery, 3D cell culture, and tissue regeneration. Conclusion Our results may provide convenient and reliable tools for detecting SAPNFs, which would be helpful for understanding their self-assembling process and exploring their applications.