Modular Design and Self-assembly of pH-Responsive Multidomain Peptides Incorporating Non-natural Ionic Amino Acids.

Stimuli-responsive peptides, particularly pH-responsive variants, hold significant promise in biomedical and technological applications by leveraging the broad pH spectrum inherent to biological environments. However, the limited number of natural pH-responsive amino acids within biologically relevant pH ranges presents challenges for designing rational pH-responsive peptide assemblies. In our study, we introduce a novel approach by incorporating a library of non-natural amino acids featuring chemically diverse tertiary amine side chains. Hydrophobic and ionic properties of these non-natural amino acids facilitate their incorporation into the assembly domain when uncharged, and electrostatic repulsion promotes disassembly under lower pH conditions. Furthermore, we observed a direct relationship between the number of substitutions and the hydrophobicity of these amino acids, influencing their pH-responsive properties and enabling rational design based on desired transitional pH ranges. The structure-activity relationship of these pH-responsive peptides was evaluated by assessing their antimicrobial properties, as their antimicrobial activity is triggered by the disassembly of peptides to release active monomers. This approach not only enhances the specificity and controllability of pH responsiveness but also broadens the scope of peptide materials in biomedical and technological applications.

In Situ Synthesis and Self-Assembly of Peptide-PEG Conjugates: A Facile Method for the Construction of Fibrous Hydrogels.

Peptide-based hydrogels have gained considerable attention as a compelling platform for various biomedical applications in recent years. Their attractiveness stems from their ability to seamlessly integrate diverse properties, such as biocompatibility, biodegradability, easily adjustable hydrophilicity/hydrophobicity, and other functionalities. However, a significant drawback is that most of the functional self-assembling peptides cannot form robust hydrogels suitable for biological applications. In this study, we present the synthesis of novel peptide-PEG conjugates and explore their comprehensive hydrogel properties. The hydrogel comprises double networks, with the first network formed through the self-assembly of peptides to create a ß-sheet secondary structure. The second network is established through covalent bond formation via N-hydroxysuccinimide chemistry between peptides and a 4-arm PEG to form a covalently linked network. Importantly, our findings reveal that this hydrogel formation method can be applied to other peptides containing lysine-rich sequences. Upon encapsulation of the hydrogel with antimicrobial peptides, the hydrogel retained high bacterial killing efficiency while showing minimum cytotoxicity toward mammalian cells. We hope that this method opens new avenues for the development of a novel class of peptide-polymer hydrogel materials with enhanced performance in biomedical contexts, particularly in reducing the potential for infection in applications of tissue regeneration and drug delivery.

A novel nanocomposite drug delivery system for SARS-CoV-2 infections.

To develop an inhalable drug delivery system, we synthesized poly (lactic-co-glycolic acid) nanoparticles with Remdesivir (RDV NPs) as an antiviral agent against SARS-CoV-2 replication and formulated Remdesivir-loaded nanocomposites (RDV NCs) via coating of RDV NPs with novel supramolecular cell-penetrating peptide nanofibers (NFs) to enhance cellular uptake and intracellular drug delivery. RDV NPs and RDV NCs were characterized using variou techniques, including Transmission Electron Microscopy (TEM), Dynamic Light Scattering (DLS), and fluorescent microscopy. The cytotoxicity of RDV NCs was assessed in Vero E6 cells and primary human lung epithelial cells, with no significant cytotoxicity observed up to 1000 µg mL-1 and 48 h. RDV NCs were spherically shaped with a size range of 200-300 nm and a zeta potential of ∼+31 mV as well as indicating the presence of coated nanofibers. Reverse Transcription-quantitative Polymerase Chain Reaction (RT-qPCR), immunofluorescence and plaque assays of SARS-CoV-2 infected Vero E6 treated with RDV NCs showed significantly higher antiviral activities compared to those of free drug and uncoated RDV NPs. RDV NCs exhibited high antiviral activity against SARS-CoV-2, and the nanocomposite platform has the potential to be developed into an inhalable drug delivery system for other viral infections in the lungs.

Self-assembling Peptides with Internal Ionizable Unnatural Amino Acids: A General Approach to pH-responsive Peptide Materials.

Self-assembled peptides are an emerging family of biomaterials that show great promise for a range of biomedical and biotechnological applications. Introducing and tuning the pH-responsiveness of the assembly is highly desirable for improving their biological activities. Inspired by proteins with internal ionizable residues, we report a simple but effective approach to constructing pH-responsive peptide assembly containing unnatural ionic amino acids with an aliphatic tertiary amine side chain. Through a combined experimental and computational investigation, we demonstrate that these residues can be accommodated and stabilized within the internal hydrophobic compartment of the peptide assembly. The hydrophobic microenvironment shifts their pKa significantly from a basic pH typically found for free amines to a more biologically relevant pH in the weakly acidic range. The pH-induced ionization and ionization-dependent self-assembly and disassembly are thoroughly investigated and correlated with the biological activity of the assembly. This new approach has unique advantages in tuning the pH-responsiveness of self-assembled peptides across a large pH range in a complex biological environment. We anticipate the ionizable amino acids developed here can be widely applicable to the synthesis and self-assembly of many amphiphilic peptides with endowed pH-responsive properties to enhance their biological activities toward applications ranging from targeted therapeutic delivery to proton transport.