Exploring the Structural Properties of alpha,gamma-Hybrid Peptide Foldamers to Design Antimicrobials, Abeta42-Oligomers Aggregation Inhibitors and Nanotubes

Abstract

The structure and functions of proteins have inspired the design of new foldamers from organic templates and unnatural amino acid building blocks. In comparison to the foldamers derived from homogenous sequences of beta- and gamma-amino acid building blocks, heterogeneous sequences composed of alpha-, beta- and gamma-amino acids offered a variety of H-bonded protein secondary structure mimetics. Among the mixed peptide sequences, alpha, gamma- and alpha, alpha, gamma-hybrid peptides have been shown to adopt stable 12- and 10/12-helical conformations. We have explored the combination of alpha- and gamma-amino acid to design different types of protein secondary structure mimetics. The present work is mainly focusing on the design of alpha, gamma-hybrid peptides as A42 aggregation inhibitors, proteolytically stable peptide antibiotics, novel beta-double helix, and hybrid peptide nanotubes to measure protein-small molecule interactions. The designed amphiphilic cationic alpha, alpha, gamma-helical peptides displayed potent broad-spectrum antimicrobial activity (AMA) across the panel of Gram-positive, and Gram-negative bacteria including MRSA and VRSA. The results of Time Kill Analysis, Post Antibiotic Effect, Skin Infection Model, and other biophysical assays suggested that these alpha, alpha, gamma-helical peptides efficiently control the growth of the bacteria through instant membrane depolarization. In addition, we have also designed cationic amphiphilic peptides and alpha, alpha, gamma -hybrid peptides as A42 aggregation inhibitors. Among various Abeta-oligomers, Abeta1-42 (Abeta42) is known to be highly aggregation-prone and also toxic. The designed peptides not only inhibit the soluble Abeta42 into aggregated fibrils but also disintegrate matured fibrils of Abeta42 into smaller assemblies. The biophysical analysis comprising the Thioflavin T assay, CD, 2D-NMR HSQC, DLS, and ELISA assays revealed that upon interaction the peptide induces a conformational change in the Abeta42. Further, the cell assay experiments revealed a significant reduction in the toxicity of the Abeta42 in the presence of cationic peptides. We further noticed that hydrophobic alpha, gamma -hybrid peptide helical foldamers spontaneously self-assembled into nanotubes and we explored these nanotubes for label-free detection of small molecule-protein interactions. The small molecules were immobilized on the surface of the functionalized nanotubes and measured the protein interactions using nanoscale capacitance and impedance spectroscopy. In comparison to the 12-helix structures of 1:1 alpha, gamma -hybrid peptides, we have shown the novel beta-double helix structures from 1:1 alternating alpha- and alpha, beta-unsaturated gamma-amino acids. The 2D NMR analysis and molecular simulations studies revealed that the peptides adopted stable anti-parallel beta-double helix conformations in solution. Overall, we have demonstrated the antimicrobial activities including drug resistant bacteria, inhibition of the aggregation of Abeta42 of Alzheimer’s disease, nanotubes as means to detect protein-small molecule interactions without labels, and novel beta-double helix structures from the mixed alpha, gamma -hybrid peptides.