Peptide Building Blocks Research Articles

Controlled peptide coated nanostructures via the self-assembly of functional peptide building blocks

To realize the self-assembly of an arginine-rich peptide sequence of R8GRGDS with tumor-targeting and membrane-penetrating functions, a hydrophobic aliphatic tail (stearic acid, C18) was coupled to its N-terminus to construct surfactant-like peptide-amphiphiles (SLPAs). Through increasing the number of C18 tails to enhance the hydrophobic interactions, SLPA2 with two C18 tails and SLPA3 with four C18 tails can self-assemble into different nanostructures, including spherical micelles, nanorods and nanofibers in aqueous solution. Because the self-assembly of SLPA2 and SLPA3 is mainly driven by the hydrophobic interactions among the C18 tails, the random-coil conformation of the functional R8GRGDS sequence does not change during self-assembly and the resulting self-assembled nanostructures can be seen as a functional R8GRGDS sequence coated architecture. When using the self-assembled micelles of SLPA3 to load the anti-tumor drug of doxorubicin (DOX) and incubating with HeLa or COS-7 cells, the DOX loaded micelles can efficiently use the tumor-targeting and membrane-penetrating functions of their surface coated R8GRGDS sequences to deliver the drug into HeLa cells. The strategy reported here presents potential for the construction of biocompatible peptide-based biomaterials with favorable bioactivity.

A modular synthetic platform for the construction of protein-based supramolecular polymers via coiled-coil self-assembly

Programmed biomolecular self-assembly is a powerful method for the construction of designed supramolecular materials. Here, we show that branched ∼8 kDa subunits composed of α-helical coiled-coil protein domains cross-linked at their midpoints by an organic bridging group can form defined supramolecular polymer assemblies in aqueous solution. The subunits are accessible by a modular and convergent synthetic route based on the chemoselective ligation of unprotected peptide building blocks. Systematic characterization of solution folding and self-assembly demonstrate that the properties of the assemblies (e.g., chain length, hydrodynamic radius) are sensitive to both the structure of the organic bridge and the sequence of the protein domain. These findings raise the possibility of the rational design of a family of supramolecular materials with tunable properties based on well-established sequence–folding relationships in coiled-coil proteins.

Optimizing the refolding conditions of self-assembling polypeptide nanoparticles that serve as repetitive antigen display systems

Nanoparticles show great promise as potent vaccine candidates since they are readily taken up by the antigen presenting cells of the immune system. The particle size and the density of the B cell epitopes on the surface of the particles greatly influences the strength of the humoral immune response. We have developed a novel type of nanoparticle composed of peptide building blocks (Raman et al., 2006) and have used such particles to design vaccines against malaria and SARS (Kaba et al., 2009, Pimentel et al., 2009). Here we investigate the biophysical properties and the refolding conditions of a prototype of these self-assembling polypeptide nanoparticles (SAPNs). SAPNs are formed from a peptide containing a pentameric and a trimeric coiled-coil domain. At near physiological conditions the peptide self-assembles into about 27 nm, roughly spherical SAPNs. The average size of the SAPNs increases with the salt concentration. The optimal pH for their formation is between 7.5 and 8.5, while aggregation occurs at lower and higher values. A glycerol concentration of about 5% v/v is required for the formation of SAPNs with regular spherical shapes. These studies will help to optimize the immunological properties of SAPNs.

ChemInform Abstract: Synthesis and Properties of MIDA Boronate Containing Aromatic Amino Acids: New Peptide Building Blocks.

AbstractNegishi‐coupling of the iodoalanines (R)‐ and (S)‐(I) with boronates (II) allows formation of the amino acid derivatives (R)‐ and (S)‐(III) in enantiomerically pure form.

Self‐Assembly of Hexagonal Peptide Microtubes and Their Optical Waveguiding

Self-assembly of biological building blocks including proteins and peptides is attracting increasing attention due to the versatility for bottom-up fabrication, biocompatibility and biodegradability, and a wide range of applications in biology and in nanotechnology. [ 1 ] In the self-assembly process the precise control of supramolecular architectures is achieved through synergistic effects of some weak non-covalent interactions such as hydrogen bonds, electrostatic interaction, π – π stacking, hydrophobic forces, nonspecifi c van der Waals forces, and chiral dipole–dipole interactions. [ 2 ] These weak forces, in particular hydrogen bonds and hydrophobic interactions, are ubiquitous in biological systems. As a whole they govern the molecular selfassembly into superior and ordered structures. By learning from nature, one can purposefully devise and synthesize artifi cial building blocks amenable to self-assembly into superstructures, which are driven by the cooperative interactions of such weak forces. With respect to other biological building blocks, peptides have been of great interest as a strategy in material fabrication owing to their structural simplicity and tunability, functional versatility, cost effectiveness, and widespread applications. As one of the simplest peptide building blocks, diphenylalanine (L-PheL-Phe, FF), a core recognition motif of Alzheimer’s β -amyloid polypeptide, [ 3 ] has been proven to be an exciting candidate towards bottom-up fabrication. FF-based building blocks are able to self-assemble into a variety of nano structures including nanotubes, spheric vesicles, nanofi brils, nanowires, and ordered molecular chains. [ 1c ] These well-defi ned nanostructures can be regarded as an assembly of molecules into supramolecular systems, however, organization of these into crystalline materials with spatial dimensions remains a formidable challenge. We report here that the FF building block is capable of selfassembling into robust hexagonal crystalline microtubes as a result of confi ned organization of nanoscale tubular structures at long range during slow crystallization or aggregation. In biological systems, the oriented organization or alignment of substructures is a common protocol to achieve functions. For

Honeycomb Self‐Assembled Peptide Scaffolds by the Breath Figure Method

The self-assembly of molecules into desired architectures is currently a challenging subject for the development of supramolecular chemistry. Here we present a facile "breath figure" assembly process through the use of the self-assembled peptide building block diphenylalanine (L-Phe-L-Phe, FF). Macroporous honeycomb scaffolds were fabricated, and average pore size could be regulated, from (1.00±0.18) μm to (2.12±0.47) μm, through the use of different air speeds. It is indicated that the honeycomb formation is humidity-, solvent-, concentration-, and substrate-dependent. Moreover, water molecules introduced from "breath figure" intervene in the formation of hydrogen bonds during FF molecular self-assembly, which results in a hydrogen bond configuration transition from antiparallel β sheet to parallel β sheet. Meanwhile, as a result of the higher polarity of water molecules, the FF molecular array is transformed from laminar stacking into a hexagonal structure. These findings not only elucidate the FF molecule self-assembly process, but also strongly support the mechanism of breath figure array formation. Finally, human embryo skin fibroblast (ESF) culture experiments suggest that FF honeycomb scaffolds are an attractive biomaterial for growth of adherent cells with great potential applications in tissue engineering.

Synthesis and properties of MIDA boronate containing aromatic amino acids: New peptide building blocks

Herein, we report the synthesis of novel phenylalanine and tyrosine derivatives containing a N-methyliminodiacetic acid boronate group. These compounds can be prepared enantiomerically pure, they are stable to column chromatography and they can be stored in air for two months without degradation occurring. This new class of boronate containing aromatic amino acids has potential applications in both peptide chemistry and natural product synthesis.

Micelle to fibre biocatalytic supramolecular transformation of an aromatic peptide amphiphile

We use a range of spectroscopic methods to provide mechanistic insight into a phosphatase-driven supramolecular transformation whereby an amphiphilic peptide building block, upon dephosphorylation, switches from a solution-phase, micellar structure to a gel-phase, chiral uni-directional nanofibre morphology.

Toroidal β-barrels from self-assembling β-sheet peptides

Controlling nanostructural morphologies has been the subject of intensive research as nanostructures can display significantly different bioactivities depending on their physicochemical parameters. Toroidal proteins perform numerous important functions in biological systems. Recent studies show that toroidal nanostructures can be constructed by rationally designed self-assembling peptide building blocks.

ChemInform Abstract: Self‐Assembly and Application of Diphenylalanine‐Based Nanostructures

AbstractReview: 66 refs.

ChemInform Abstract: Diastereoselective Synthesis of C-Glycosylated Amino Acids with Lactams as Peptide Building Blocks.

ChemInform Abstract: Diastereoselective Synthesis of C-Glycosylated Amino Acids with Lactams as Peptide Building Blocks.

Copper-Catalyzed N-Arylation of Semicarbazones for the Synthesis of Aza-Arylglycine-Containing Aza-Peptides

Parallel synthesis of 13 aza-arylglycine peptides, based on the hexapeptide sequence of Growth Hormone Releasing Peptide-6 (GHRP-6), was accomplished via selective N-arylation of a semicarbazone peptide building block anchored on Rink amide resin. Aza-peptides possessing aza-indolylglycine and aza-imidazoylglycine residues were obtained through use of the corresponding heteroaryl iodides, yielding, respectively, aza-Trp and aza-His peptidomimics. CD spectroscopy indicated the propensity for aza-peptides, containing aza-arylglycines at the Trp(4) position of the GHRP-6 sequence, to adopt beta-turns.

Constructing biomaterials using self-assembling peptide building blocks

Molecular self-assembly is ubiquitous in nature and has recently emerged as a new bottom-up approach in constructing biomaterials. Synthetic peptides assemble through specific molecular recognition and form diverse nanostructures. The resulting versatile peptide self-assemblies may be used in a wide range of biological and medical applications. Examples of two self-assembling peptide systems are presented and techniques for self-assembly control are discussed.

Self-assembly and application of diphenylalanine-based nanostructures

Micro- and nanostructures fabricated from biological building blocks have attracted tremendous attention owing to their potential for application in biology and in nanotechnology. Many biomolecules, including peptides and proteins, can interact and self-assemble into highly ordered supramolecular architectures with functionality. By imitating the processes where biological peptides or proteins are assembled in nature, one can delicately design and synthesize various peptide building blocks composed of several to dozens of amino acids for the creation of biomimetic or bioinspired nanostructured materials. This tutorial review aims to introduce a new kind of peptide building block, the diphenylalanine motif, extracted with inspiration of a pathogenic process towards molecular self-assembly. We highlight recent and current advances in fabrication and application of diphenylalanine-based peptide nanomaterials. We also highlight the preparation of such peptide-based nanostructures as nanotubes, spherical vesicles, nanofibrils, nanowires and hybrids through self-assembly, the improvement of their properties and the extension of their applications.

Structures of self-assembled amphiphilic peptide-heterodimers: effects of concentration, pH, temperature and ionic strength

The amphiphilic double-tail peptides AXG were studied regarding secondary structure and self-assembly in aqueous solution. The two tails A = Ala6 and G = Gly6 are connected by a central pair X of hydrophilic residues, X being two aspartic acids in ADG, two lysines in AKG and two arginines in ARG. The peptide AD (Ala6Asp) served as a single-tail reference. The secondary structure of the four peptides was characterized by circular dichroism spectroscopy under a wide range of peptide concentrations (0.01–0.8 mM), temperatures (20–98 °C), pHs (4–9.5) and ionic strengths. In salt-free water both ADG and AD form a β-sheet type of structure at high concentration, low pH and low temperature, in a peptide–peptide driven assembly of individual peptides. The transition has a two-state character for ADG but not for AD, which indicates that the added tail in ADG makes the assembly more cooperative. By comparison the secondary structures of AKG and ARG are comparatively stable over the large range of conditions covered. According to dynamic light scattering the two-tail peptides form supra-molecular aggregates in water, but high-resolution AFM-imaging indicate that ordered (self-assembled) structures are only formed when salt (0.1 M NaCl) is added. Since the CD-studies indicate that the NaCl has only a minor effect on the peptide secondary structure we propose that the main role of the added salt is to screen the electrostatic repulsion between the peptide building blocks. According to the AFM images ADG and AKG support a correlation between nanofibers and a β-sheet or unordered secondary structure, whereas ARG forms fibers in spite of lacking β-sheet structure. Since the AKG and ARG double-tail peptides self-assemble into distinct nanostructures while their secondary structures are resistant to environment factors, these new peptides show potential as robust building blocks for nano-materials in various medical and nanobiotechnical applications.

Amyloid-Like Self-Assembly of Peptide Sequences from the Adenovirus Fiber Shaft: Insights from Molecular Dynamics Simulations

The self-assembly of peptides and proteins into nanostructures is related to the fundamental problems of protein folding and misfolding and has potential applications in medicine, materials science and nanotechnology. Natural peptides, corresponding to sequence repeats from self-assembling proteins, may constitute elementary building blocks of such nanostructures. In this work, we study by implicit-solvent replica-exchange simulations the self-assembly of two amyloidogenic sequences derived from the naturally occurring fiber shaft of the adenovirus, the octapeptide NSGAITIG (asparagine-serine-glycine-alanine-isoleucine-threonine-isoleucine-glycine) and its hexapeptide counterpart, GAITIG. In accordance with their amyloidogenic capacity, both peptides form readily intermolecular beta-sheets, stabilized by extensive main- and side-chain contacts involving the C-terminal moieties (segments 3-8 and 2-6, respectively). The structural and energetic properties of these sheets are analyzed extensively. The N-terminal residues Asn1 and Ser2 of the octapeptide remain disordered in the sheets, suggesting that these residues are exposed at the exterior of the fibrils and accessible. On the basis of insight provided by the simulations, cysteine residues were recently substituted at positions 1 and 2 of NSGAITIG; the newly designed peptides maintain their amyloidogenic properties and can bind to silver, gold and platinum nanoparticles [Kasotakis et al. Biopolymers 2009, 92, 164-172]. Computational investigation can identify suitable positions for rational modification of peptide building blocks, aiming at the fabrication of novel biomaterials.

High stability of self‐assembled peptide nanowires against thermal, chemical, and proteolytic attacks

Understanding the self-assembly of peptides into ordered nanostructures is recently getting much attention since it can provide an alternative route for fabricating novel bio-inspired materials. In order to realize the potential of the peptide-based nanofabrication technology, however, more information is needed regarding the integrity or stability of peptide nanostructures under the process conditions encountered in their applications. In this study, we investigated the stability of self-assembled peptide nanowires (PNWs) and nanotubes (PNTs) against thermal, chemical, proteolytic attacks, and their conformational changes upon heat treatment. PNWs and PNTs were grown by the self-assembly of diphenylalanine (Phe-Phe), a peptide building block, on solid substrates at different chemical atmospheres and temperatures. The incubation of diphenylalanine under aniline vapor at 150 degrees C led to the formation of PNWs, while its incubation with water vapor at 25 degrees C produced PNTs. We analyzed the stability of peptide nanostructures using multiple tools, such as electron microscopy, thermal analysis tools, circular dichroism, and Fourier-transform infrared spectroscopy. Our results show that PNWs are highly stable up to 200 degrees C and remain unchanged when incubated in aqueous solutions (from pH 1 to 14) or in various chemical solvents (from polar to non-polar). In contrast, PNTs started to disintegrate even at 100 degrees C and underwent a conformational change at an elevated temperature. When we further studied their resistance to a proteolytic environment, we discovered that PNWs kept their initial structure while PNTs fully disintegrated. We found that the high stability of PNWs originates from their predominant beta-sheet conformation and the conformational change of diphenylalanine nanostructures. Our study suggests that self-assembled PNWs are suitable for future nano-scale applications requiring harsh processing conditions.

Functional Ubiquitin Conjugates with Lysine-ε-Amino-Specific Linkage by Thioether Ligation of Cysteinyl-Ubiquitin Peptide Building Blocks

The modification of ubiquitin to defined oligo-ubiquitinated conjugates has received considerable interest due to the finding that isomeric oligo-ubiquitin conjugates exhibit distinct differences in their biochemical functions, depending on the specific lysine-epsilon-amino linkage used for conjugate formation. Here, we report the design and development of a thioether linkage-based approach for the synthesis of oligo-ubiquitin conjugates with lysine-specific branching by thioether ligation of a linear ubiquitin peptide containing a C-terminal cysteine residue as the "donor" component, with a corresponding lysine-epsilon-amino-branched haloacyl-activated ubiquitin "acceptor" peptide. This approach was successfully used for the synthesis of a lysine-63-linked diubiquitin conjugate by ligation of the modified ubiquitin(1-52)-Cys- donor peptide to the N-terminal Arg-54 residue of the branched Lys-63-linked acceptor peptide, ubiquitin(54-76)(2). Advantages of the present approach are as follows: (i) the conjugation reaction is performed in solution using suitable preformed donor ubiquitin peptides with a C-terminal Cys residue, and (ii) different corresponding N-chloroacetylated ubiquitin acceptor peptides containing the branched Lys residue are employed, providing broad applicability to the preparation of isomeric oligo-ubiquitin conjugates. The Lys-63-diubiquitin conjugate 7 described here was purified by semipreparative HPLC, and its structure and homogeneity ascertained by HPLC and high-resolution MALDI and electrospray-mass spectrometry. CD spectra and molecular modeling indicate a conformationally stable structure of the conjugate with spatial separation of the ubiquitin parts of the Lys-63 linkage. Moreover, the activity of the thioether-linked diubiquitin conjugate was ascertained by in vitro autoubiquitination assay. These results indicate the feasibility of this approach for the preparation of functional oligo-ubiquitin conjugates.

Chiral and Achiral Fundamental Conformational Building Units of β-Peptides: A Matrix Isolation Conformational Study on Ac-β-HGly-NHMe and Ac-β-HAla-NHMe

The infrared spectra of two model beta-peptides, N-acetyl-3-aminopropionic acid-N'-methylamide (Ac-beta-HGly-NHMe) and N-acetyl-3-aminobutanoic acid-N'-methylamide (Ac-beta-HAla-NHMe), have been recorded in low-temperature Ar and Kr matrixes. The spectra were assigned by the help of electronic structure calculations. The analysis of spectra, in line with the theoretical predictions, revealed that both biocompatible peptide building blocks have a single dominant backbone conformer. Besides this prevalent conformer, which has a six-membered H-bonded pseudoring, conformers with eight-membered H-bonded rings are also observed but in a significantly smaller amount. The calculated conformer distribution is consistent with the experimental findings. The present work along with other recent results supports the concept that the backbone conformation of longer biopolymers, such as alpha- and beta-peptides, can be deciphered using the conformers of their structural building blocks. In this respect, our conformational study on the simplest models for beta-peptides both by IR spectroscopic experiments and quantum chemical studies has significance for the better design and understanding of the backbone conformations of larger beta-peptides with biomedical potential. The present conformational analysis suggests that although beta-peptides, having an "extra" backbone torsion and hence more conformational freedom, should be more flexible than alpha-peptides, fewer backbone conformers are viable based on their relative energies. Thus, from a larger conformational arsenal, only a lower number of backbone conformers can emerge, which possibly had a fundamental effect on their applicability during prebiotic evolution.

Designed amphiphilic peptide forms stable nanoweb, slowly releases encapsulated hydrophobic drug, and accelerates animal hemostasis

How do you design a peptide building block to make 2-dimentional nanowebs and 3-dimensional fibrous mats? This question has not been addressed with peptide self-assembling nanomaterials. This article describes a designed 9-residue peptide, N-Pro-Ser-Phe-Cys-Phe-Lys-Phe-Glu-Pro-C, which creates a strong fishnet-like nanostructure depending on the peptide concentrations and mechanical disruptions. This peptide is intramolecularly amphiphilic because of a single pair of ionic residues, Lys and Glu, at one end and nonionic residues, Phe, Cys, and Phe, at the other end. Circular dichroism and Fourier transform infrared spectroscopy analysis demonstrated that this peptide adopts stable beta-turn and beta-sheet structures and self-assembles into hierarchically arranged supramolecular aggregates in a concentration-dependent fashion, demonstrated by atomic force microscopy and electron microscopy. At high concentrations, the peptide dominantly self-assembled into globular aggregates that were extensively connected with each other to form "beads-on-a-thread" type nanofibers. These long nanofibers were extensively branched and overlapped to form a self-healing peptide hydrogel consisting of >99% water. This peptide can encapsulate the hydrophobic model drug pyrene and slowly release pyrene from coated microcrystals to liposomes. It can effectively stop animal bleeding within 30 s. We proposed a plausible model to interpret the intramolecular amphiphilic self-assembly process and suggest its importance for the future development of new biomaterials for drug delivery and regenerative medicine.