Fluorescence Correlation Spectroscopy Analysis of Effect of Molecular Crowding on Self-Assembly of β-Annulus Peptide into Artificial Viral Capsid

Recent progress in the de novo design of self-assembling peptides has enabled the construction of peptide-based viral capsids. Previously, we demonstrated that 24-mer β-annulus peptides from tomato bushy stunt virus spontaneously self-assemble into an artificial viral capsid. Here we propose to use the artificial viral capsid through the self-assembly of β-annulus peptide as a simple model to analyze the effect of molecular crowding environment on the formation process of viral capsid. Artificial viral capsids formed by co-assembly of fluorescent-labelled and unmodified β-annulus peptides in dilute aqueous solutions and under molecular crowding conditions were analyzed using fluorescence correlation spectroscopy (FCS). The apparent particle size and the dissociation constant (K_d) of the assemblies decreased with increasing concentration of the molecular crowding agent, i.e., polyethylene glycol (PEG). This is the first successful in situ analysis of self-assembling process of artificial viral capsid under molecular crowding conditions.     

Enhancing the Cellular Delivery of Nanoparticles Using Lipo‐Oligoarginine Peptides

Nanoparticles have great potential as nanotherapeutics, delivery vectors, and molecular imaging agents due to their flexible properties. Although intracellular and nuclear delivery of nanoparticles is desirable for therapeutic applications, it remains a challenge. Cell penetrating peptides (CPPs) are a powerful tool for the intracellular delivery of various cargoes. Here it is reported that functionalization of nanoparticles with a myristoylated oligoarginine CPP promotes cellular uptake without increased toxicity. It is evident that the myristoylated CPP is much more effective in transporting nanoparticles than the unmodified CPPs.     

Peptide‐functionalized reduced graphene oxide as a bioactive mechanically robust tissue regeneration scaffold

Bioactive, synthetic materials represent next‐generation composites for tissue regeneration. Design of contemporary materials attempts to recapitulate the complexities of native tissue; however, few successfully mimic the order in nature. Recently, graphene oxide (GO ) has emerged as a scaffold due to its potential for bioactive functionalization and long‐range order instilled by the self‐assembly of graphene sheets. Chemical reduction of GO results in a more compatible material with enhanced properties but compromises the ability to functionalize the graphenic backbone. However, using Johnson–Claisen rearrangement chemistry, functionalization is achieved that is not liable to reduction. From reduced Claisen graphene, we polymerized short homopeptides from α ‐amino acid N ‐carboxyanhydride monomers of glutamate and lysine to result in functionalized graphenes (pGlu‐rCG and pLys‐rCG ) that are cytocompatible, degradable, and bioactive. Exposure to NIH‐3T3 fibroblasts and RAW 264.7 macrophages revealed that the materials are cytocompatible and do not alter important sub‐cellular compartments. Powders were hot pressed to form mechanically stiff (E ′: 41 and 49 MPa ), strong (UCS : 480 and 140 MPa ), and tough (U _T: 2898 and 584 J m^?3 × 10⁴) three‐dimensional constructs (pGlu‐rCG and pLys‐rCG, respectively). Overall, we report a robust chemistry and processing strategy for facile bioactive functionalization of compatible, reduced Claisen graphene for three‐dimensional biomedical applications. © 2017 Society of Chemical Industry     

One-Step Chemical Synthesis of Native Met1-Linked Poly-Ubiquitin Chains

The enzyme-mediated construction of poly-ubiquitin (Ub) chains on target proteins leads to a variety of cellular responses and is involved in processes ranging from protein degradation to cell cycle control and immune responses. This complex post-translational modification system is under intense investigation, but generation of specific Ub chains and tools made thereof is not always trivial. We discovered that native methionine-1-linked polymeric ubiquitin chains can be constructed in a single chemical reaction. We validate correct folding and regioselective attachment of such chains using linkage specific proteases and further demonstrate that these poly-Ub chains can be converted into thioesters by the activating E1-enzyme. Subsequent ligation reactions using these in situ prepared thioesters leads to poly-ubiquitinated peptides.     

Order/Disorder in Protein and Peptide-Based Biomaterials

Nature utilizes both order and disorder (or controlled disorder) to achieve exceptional materials properties and functions, while synthetic supramolecular materials mostly exploit just supramolecular order, thus limiting the structural diversity, responsiveness and consequent adaptive functions that can be accessed. Herein, we review the emerging field of supramolecular biomaterials where disorder and order deliberately co-exist, and can be dynamically regulated by considering both entropic and enthalpic factors in design. We focus on sequence-structure relationships that govern the (cooperative) assembly pathways of protein and peptide building blocks in these materials. Increasingly, there is an interest in introducing dynamic features in protein and peptide-based structures, such as the remarkable thermo-responsiveness and exceptional mechanical properties of elastin materials. Simultaneously, advances in the field of intrinsically disordered proteins (IDPs) give new insights about their involvement in intracellular liquid-liquid phase separation and formation of disordered, dynamic coacervate structures. These have inspired efforts to design biomaterials with similar dynamic properties. These hybrid ordered/disordered materials employ a combination of intramolecular and supramolecular order/disorder features for construction of assemblies that are dynamically reconfigurable. The assembly of these dynamic structures is mainly entropy-driven, relying on electrostatic and hydrophobic interactions and is mediated in part through the adopted (unstructured) protein conformation or by introducing an oppositely charged guest for peptide building blocks. Examples include design of protein building blocks composed of disordered repeat sequences of elastin-like polypeptides in combination with ordered regions that adopt a secondary structure, the co-assembly of proteins with peptide amphiphiles to achieve reconfigurable, yet highly stable membranes or tyrosine-containing tripeptides with sequence-controlled order/disorder that upon enzymatic oxidation give rise to melanin-like polymeric pigments with customizable properties. The resulting hybrid materials with controlled disorder can be metastable, and sensitive to various external stimuli giving rise to insights that are especially attractive for the design of responsive and adaptive materials.     

Detailed Insight into the Interaction of Bicyclic Somatostatin Analogue with Cu(II) Ions

Somatostatin analogues are useful pharmaceuticals in peptide receptor radionuclide therapy. In previous studies, we analyzed a new bicyclic somatostatin analogue (BCS) in connection with Cu(II) ions. Two characteristic sites were present in the peptide chain: the receptor- and the metal-binding site. We have already shown that this ligand can form very stable imidazole complexes with the metal ion. In this work, our aim was to characterize the intramolecular interaction that occurs in the peptide molecule. Therefore, we analyzed the coordination abilities of two cyclic ligands, i.e., P1 only with the metal binding site and P2 with both sites, but without the disulfide bond. Furthermore, we used magnetic circular dichroism (MCD) spectroscopy to better understand the coordination process. We applied this method to analyze spectra of P1, P2, and BCS, which we have described previously. Additionally, we analyzed the MCD spectra of P3 ligand, which has only the receptor binding site in its structure. We have unequivocally shown that the presence of the Phe-Trp-Lys-Thr motif and the disulfide bond significantly increases the metal binding efficiency.     

Antioxidant and antibacterial activities of peptide fractions from flaxseed protein hydrolysed by protease from Bacillus altitudinis HK02

Flaxseed protein (FP) hydrolysates by crude protease of strain Bacillus altitudinis HK02 were further separated into five fractions by ultrafiltration membranes with a molecular weight cut‐off of 10, 5, 3 and 1 kDa for the analysis of antioxidant activities and antibacterial ability. The results demonstrated that the fraction of 1‐ to 3‐kDa peptides exhibited higher antioxidant activities on the free radical‐scavenging ability and the reducing power than those of Vit C, Vit E, BHA and other fractions. The fraction with a low molecular weight (<1 kDa) of peptides demonstrated the highest ferrous ion‐chelating ability and a higher inhibition of lipid peroxidation than BHA and other fractions. Moreover, it also exhibited the best growth inhibition of Pseudomonas aeruginosa and Escherichia coli. The results, including the concentration‐dependent effect of peptides fractions, demonstrated that it is feasible to derive functional ingredients of natural antioxidants along with antimicrobial activity from FPs hydrolysed by protease from B. altitudinis HK02.     

Recent Advances in Late-Stage Construction of Stapled Peptides via C−H Activation

Stapled peptides have been widely applied in many fields, including pharmaceutical chemistry, diagnostic reagents, and materials science. However, most traditional stapled peptide preparation methods rely on prefunctionalizations, which limit the diversity of stapled peptides. Recently, the emergence of late-stage transition metal-catalyzed C−H activation in amino acids and peptides has attracted wide interest due to its robustness and applicability for peptide stapling. In this review, we summarize the methods for late-stage construction of stapled peptides via transition metal-catalyzed C−H activation.     

Comparisons of β-Hairpin Propensity Among Peptides with Homochiral or Heterochiral Strands

Assemblies of racemic β-sheet-forming peptides have attracted attention for biomedical applications because racemic forms of peptides can self-associate more avidly than do single enantiomers. In 1953, Pauling and Corey proposed “rippled β-sheet” modes of H-bond-mediated interstrand assembly for alternating L- and D-peptide strands; this structural hypothesis was complementary to their proposal of “pleated β-sheet” assembly for L-peptides. Although no high-resolution structure has been reported for a rippled β-sheet, there is strong evidence for the occurrence of rippled β-sheets in some racemic peptide assemblies. Here we compare propensities of peptide diastereomers in aqueous solution to form a minimum increment of β-sheet in which two antiparallel strands associate. β-Hairpin folding is observed for homochiral peptides with aligned nonpolar side chains, but no β-hairpin population can be detected for diastereomers in which one strand contains L residues and the other contains D residues. These observations suggest that rippled β-sheet assemblies are stabilized by interactions between β-sheet layers rather than interactions within these layers.