The uncovering of single peptides derived from food sources that can form hydrogels is of great relevance for several applications. However, identifying single peptide hydrogels from food is a daunting task given the complex nature of the food systems. The proof of concept of the applicability of TANGO, a statistical mechanical -based algorithm that predicts the b -aggregate propensity of peptides, as a tool to uncover peptides derived from milk that can form hydrogels is reported. Using TANGO in conjunction with a set of defined criteria we discovered that from a group of thirteen peptides derived from milk proteins, seven formed hydrogels at a concentration of 2 wt% and pH 7 at room temperature. Three more peptides formed aggregates and appeared to go through the syneresis process, and three additional peptides remained liquid under the experimental conditions. This result sets the basis of a simple methodology for unveiling peptide hydrogels from food and other natural sources. (c) 2024 The Author(s). Published by Elsevier Ltd. This is an open access article under the CC BY -NC license (http://creativecommons.org/licenses/by-nc/4.0/).

In this work the identification of peptides derived from quinoa proteins which could potentially self-assemble, and form hydrogels was carried out with TANGO, a statistical mechanical based algorithm that predicts β-aggregate propensity of peptides. Peptides with the highest aggregate propensity were subjected to gelling screening experiments from which the most promising bioactive peptide with sequence KIVLDSDDPLFGGF was selected. The self-assembling and hydrogelation properties of the C-terminal amidated peptide (KIVLDSDDPLFGGF-NH2) were studied. The effect of concentration, pH, and temperature on the secondary structure of the peptide were probed by circular dichroism (CD), while its nanostructure was studied by transmission electron microscopy (TEM) and small-angle neutron scattering (SANS). Results revealed the existence of random coil, α-helix, twisted β-sheet, and well-defined β-sheet secondary structures, with a range of nanostructures including elongated fibrils and bundles, whose proportion was dependant on the peptide concentration, pH, or temperature. The self-assembly of the peptide is demonstrated to follow established models of amyloid formation, which describe the unfolded peptide transiting from an α-helix-containing intermediate into β-sheet-rich protofibrils. The self-assembly is promoted at high concentrations, elevated temperatures, and pH values close to the peptide isoelectric point, and presumably mediated by hydrogen bond, hydrophobic and electrostatic interactions, and π-π interactions (from the F residue). At 15 mg/mL and pH 3.5, the peptide self-assembled and formed a self-supporting hydrogel exhibiting viscoelastic behaviour with G' (1 Hz) ~2300 Pa as determined by oscillatory rheology measurements. The study describes a straightforward method to monitor the self-assembly of plant protein derived peptides; further studies are needed to demonstrate the potential application of the formed hydrogels in food and biomedicine.

Re-assembled casein micelles (rCMs), were formulated in the 1970s as a model system to understand native casein micelles (nCMs) in milk. These early works allowed an understanding of the critical factors involved in the formation of rCMs, such as minerals (citrate, phosphate, and calcium), casein type (αs-, β-, and κ-casein) and the extent of their phosphorylation. rCMs were also used to understand the effect of treatments such as ethanol, high hydrostatic pressure and heating on the stability and integrity of the micelles. More recently, the applications of rCMs have been investigated, these include their use as a nanocarrier of bioactive molecules and as electrode-bound substrates to monitor chymosin activity by electrochemistry, to cite a few. Moreover, the potential to use rCMs in both food and non-food applications remains to be fully exploited. The advantage of choosing rCMs over nCMs as an encapsulant and a lucrative food ingredient is due to their more efficient preparation and being free from impurities. In this review, we report on the formulation of rCMs, their physico-chemical properties and their behavior under different physico-chemical treatments, along with the applications and challenges of rCMs in food systems and their industrial production as a dairy ingredient.

Peptide and protein selective modification at tyrosine residues has become an exploding field of research as tyrosine constitutes a robust alternative to lysine and cysteine-targeted traditional peptide/protein modification protocols. This review offers a comprehensive summary of the latest advances in tyrosine-selective cleavage, functionalization, and conjugation of peptides and proteins from the past three years. This updated overview complements the extensive body of work on site-selective modification of peptides and proteins, which holds significant relevance across various disciplines, including chemical, biological, medical, and material sciences.

A novel tyrosine hyperoxidation enabling selective peptide cleavage is reported. The scission of the N-terminal amide bond of tyrosine was achieved with Dess-Martin periodinane under mild conditions, generating a C-terminal peptide fragment bearing the unprecedented hyperoxidized tyrosine motif, 4,5,6,7-tetraoxo-1H-indole-2-carboxamide, along with an intact N-terminal peptide fragment. This reaction proceeds with high site-selectivity for tyrosine and exhibits broad substrate scope for various peptides, including those containing post-translational modifications. More importantly, this oxidative cleavage was successfully applied to enable sequencing of three naturally occurring cyclic peptides, including one depsipeptide and one lipopeptide. The linearized peptides generated from the cleavage reaction significantly simplify cyclic peptide sequencing by MS/MS, thus providing a robust tool to facilitate rapid sequence determination of diverse cyclic peptides containing tyrosine. Furthermore, the highly electrophilic nature of the hyperoxidized tyrosine unit disclosed in this work renders it an important electrophilic target for the selective bioconjugation or synthetic manipulation of peptides containing this unit.

In this work we report the synthesis of mono lipidated peptides containing a 3-mercaptopropionate linker in the N-terminus by means of a photoinitiated thiol-ene reaction (S-lipidation). We evaluate the self-assembling and hydrogelation properties of a library of mono S-lipidated peptides containing lipid chains of various lengths and demonstrate that hydrogelation was driven by a balance between the lipid chain's hydrophobicity and the peptide's facial hydrophobicity. We further postulate that a simple calculation using estimated values of log D could be used as a predictor of hydrogelation when designing similar systems. A mono S-lipidated peptide containing a short lipid chain that formed hydrogels was fully characterized and a mechanism for the peptide hydrogelation developed. Finally, we demonstrate that the presence of the thioether group in the mono S-lipidated peptide hydrogels, which is a feature lacking in conventional N-acyl lipidated systems, enables the controlled disassembly of the gel via oxidation to the sulfoxide by reactive oxygen species in accordance with a hydrophobicity-modulated strategy. Thus, we conclude that mono S-lipidated peptide hydrogels constitute a novel and simple tool for the development of tissue engineering and targeted drug delivery applications of diseases with overexpression of reactive oxygen species (e.g. degenerative and metabolic diseases, and cancers).

The use of peptide hydrogels is of growing interest in bone regeneration. Self-assembling peptides form hydrogels and can be used as injectable drug delivery matrices. Injected into the defect site, they can gel in situ, and release factors that aid bone growth. We report on the design, synthesis and characterization of three β-hairpin peptide hydrogels, and on their osteoblast cytocompatibility as well as delivery of the lactoferrin glycoprotein, a bone anabolic factor. Osteoblasts cultured in hydrogels of the peptide with sequence NH2-Leu-His-Leu-His-Leu-Lys-Leu-Lys-Val-dPro-Pro-Thr-Lys-Leu-Lys-Leu-His-Leu-His-Leu-Arg-Gly-Asp-Ser-CONH2 (H4LMAX-RGDS) increased the osteoblast cell number and the cells appeared healthy after seven days. Furthermore, we showed that H4LMAX-RGDS was capable of releasing up to 60% of lactoferrin (pre-encapsulated in the gel) over five days while retaining the rest of the glycoprotein. Thus, H4LMAX-RGDS hydrogels are cytocompatible with primary osteoblasts and capable of delivering bio-active lactoferrin that increases osteoblast cell number.

Increased water solubility and long-range intermolecular ordering have been introduced into the fluorescent organic molecule thiophene-diketopyrrolopyrrole (TDPP) via its conjugation to the octapeptide HEFISTAH, which is derived from the protein-protein β-interface of the homo-tetramer protein diaminopimelate decarboxylase. The octapeptide, and its TDPP mono- and cross-linked conjugates were synthesised using 9-fluorenylmethoxycarbonyl (Fmoc) based solid-phase peptide synthesis (SPPS). Unlike the unmodified peptide, the resulting mono-linked and cross-linked peptides showed a fibrous morphology and formed hydrogels at 4 wt% in water at neutral pH, but failed to assemble at pH 2 and pH 9. Further peptide characterization showed that the TDPP organic core enhances peptide self-assembly and that both peptides assembled into fibers with a parallel β-sheet structure. Furthermore, UV-vis spectroscopic analysis suggests that the TDPP molecules form H-type aggregates where the chromophores are likely to be co-facially packed, but rotationally and/or laterally offset from one another. This intermolecular coupling indicates that π-π stacking interactions are highly likely - a favourable sign for charge transport. The enhanced aqueous solubility and self-assembling properties of the TDPP-peptide conjugates allowed the successful preparation of thin films. Atomic force microscopy, X-ray diffraction and UV-vis spectroscopic analysis of these thin films revealed that the hybrid materials retained a fibrous morphology, β-sheet structures and strong intermolecular coupling between neighbouring TDPP molecules. These results open an exciting avenue for bio-organic materials development, through structural and electronic tuning of the TDPP core.

The effect of installing different lipid chains (C-6 , C-8 , C-10, and C-16) on the N-terminus of an octapeptide derived from the antiparallel ,B-interface of the diaminopimelate decarboxylase protein homotetramer has been investigated. Notably, the C-8 peptide conjugate assembled into wide twisted nanoribbons and formed hydrogels, which to the best of our knowledge constitutes the first example of a peptide containing an eight carbon alkyl chain that demonstrates these properties, a space typically occupied by peptide amphiphiles with long lipid chains. Furthermore, this self-assembling lipopeptide exhibited pH and temperature stability with shear thinning properties suitable for biomedical applications. Importantly, in this work the application of the polystyrene-based sorbent DiaionTM HP20SS for the simple large-scale purification of self-assembling peptides is presented as an alternative to the use of time-consuming and labor-intensive reverse-phase high-performance liquid chromatography. Statement of Significance Peptides that can self-assemble into defined nanostructures are highly attractive for many biomedical applications given their unique physical and chemical properties. It is recognized that self-assembling peptides derived from naturally occurring proteins offer an unlimited source of functionalities and structures, which are hard to uncover with designed sequences. In this study, we have investigated the effect of installing different lipids chains on the N-terminus of an octapeptide derived from the antiparallel ,B-interface of the diaminopimelate decarboxylase protein homo tetramer. We also reported the use of polymeric Diaion (R) HP20SS beads as an alternative solid support to purify self-assembling peptides. (C) 2020 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Background: The interest in peptide hydrogels of natural origin has dramatically increased given the potential applications in several fields -e.g. biomedicine and nanotechnology. Interestingly, despite the current knowledge on protein hydrolysates from food sources, which self-assemble and form gels, the extraction of single peptides that can form hydrogels from food products and/or their application in food and other areas remains poorly explored. Scope and approach: This review provides a prospective analysis of the literature on the mechanism, production, toxicity and potential applications of food derived peptide hydrogels. Key findings and conclusions: Food products can be an important source of single peptides that form hydrogels, and these can find applications in food science and other areas. However, research in this area is in its infancy and its progress is limited due in part to the lack of 1) tools that will allow one to predict peptide fragments within a food protein that can self-assemble and form gels and 2) efficient peptide purification protocols. Therefore, more research will have to be directed on these areas in conjunction with optimization of recombinant, and enzymatic/fermentation production protocols.

Wollamides are cyclic hexapeptides, recently isolated from an Australian soil Streptomyces isolate, that exhibit promising in vitro antimycobacterial activity against Mycobacterium bovis Bacille Calmette Guérin without displaying cytotoxicity against a panel of mammalian cells. Here, we report the synthesis and antimycobacterial activity of 36 new synthetic wollamides, collated with all known synthetic and natural wollamides, to reveal structure characteristics responsible for in vitro growth-inhibitory activity against Mycobacterium tuberculosis (H37Rv, H37Ra, CDC1551, HN878, and HN353).

The self-assembly of peptide nanotubes (PNTs) depends on the structure and chemistry of cyclic peptide (CP) monomers, having an impact on their properties, making the choice of their monomers and their characterization a great challenge. We synthesized for the first time a new set of eight original CP sequences of 8, 10, and 12 D,L-alpha-alternate amino acids with a controlled internal diameter from 7 to 13 A. They present various properties (e.g., diameter, global surface charge, hydrophobicity) that can open the way to new applications. Their structure and purity were determined thanks to a capillary electrophoresis coupled to electrospray ionization mass spectrometry (CE-ESI-MS) methodology developed for the first time for this purpose. The CPs were successfully separated in a basic hydro-organic background electrolyte (BGE, pH 8.0, H2O/EtOH 50:50, v/v) and analyzed in MS positive mode. The effect of CP structure on electrophoretic mobility was studied, and the mass spectra were deeply analyzed. This methodology allowed verifying their purity and the absence of linear peptide precursors as well as their stability when stored over several months. Therefore, we have developed a new CE-ESI-MS methodology for the structure and purity control of interesting potential precursors for PNTs that could be employed as nanoplatforms in diagnostics or as pseudo sieving tools for separative purposes. (C) 2016 Elsevier Inc. All rights reserved.

The development of synthetic methods to prepare conformationally constrained peptides and peptide-polyketide hybrids remain an important chemical challenge. It is known that structural rigidity correlates with the specificity, bioactivity, and stability of these peptide systems, thus rigid systems are particularly attractive leads for development of potent biopharmaceuticals. Herein we provide an overview of recent developments in the syntheses of naturally derived constrained peptides and peptide-polyketide hybrids, with a particular emphasis on those systems containing an ene-like bond.

The proposed structure of talarolide A, a cycloheptapeptide featuring a hydroxamate moiety within the peptide backbone, was successfully synthesized. An initial attempt to synthesize a linear peptide precursor containing a C-terminal N-benzyloxy glycine residue was problematic due to an unreported on-resin reduction of N-benzyloxy glycine to glycine. After repositioning the peptide cyclization point, a new linear peptide sequence was successfully prepared using Fmoc-solid-phase peptide synthesis. Subsequent solution-phase cyclization and removal of protecting groups furnished the synthetic talarolide A in good yield. Despite the mismatch of the NMR data between the synthetic talarolide A and the natural product, a detailed structural analysis using 2D NMR spectroscopy, together with re-synthesis of the same synthetic material using two additional cyclization sites, confirmed that our synthetic product has the reported structure of talarolide A.

Callyaerin A ist ein natürliches cyclisches Peptid mit dem seltenen (Z)-2,3-Diaminoacrylamid(DAA)-Motiv. Es wirkt stark inhibierend gegen M. tuberculosis, ist aber nicht toxisch in menschlichen Zellen. In ihrer Zuschrift auf S. 3693 beschreiben M. Brimble et al. die Totalsynthese von Callyaerin A und zeigen, dass dem Peptid durch die DAA-Einheit eine starre Konformation aufgezwungen wird. Diese Ergebnisse weisen den Weg für weitere Untersuchungen zum potenziellen therapeutischen Nutzen des Peptids.

The first synthesis of the anti-TB cyclic peptide callyaerin A containing a rare (Z)-2,3-diaminoacrylamide bridging motif is reported. Fmoc-formylglycine-diethylacetal was used as a masked equivalent of formylglycine in the synthesis of the linear precursor of callyaerin A. Intramolecular cyclization between the formylglycine residue and the N-terminal amine in the linear peptide precursor afforded the macrocyclic natural product callyaerin A. Synthetic callyaerin A possessed potent anti-TB activity (MIC100 = 32 μM) while its all-amide congener was inactive. Variable temperature NMR studies of both the natural product and its all amide analogue revealed the extraordinary rigidity imposed by this diaminoacrylamide unit on peptide conformation. The work reported herein pinpoints the intrinsic role that the rare (Z)-2,3-diaminoacrylamide moiety confers on peptide

The development of synthetic methods to prepare conformationally constrained peptides and peptide-polyketide hybrids remain an important chemical challenge. It is known that structural rigidity correlates with the specificity, bioactivity, and stability of these peptide systems, thus rigid systems are particularly attractive leads for development of potent biopharmaceuticals. Herein we provide an overview of recent developments in the syntheses of naturally derived constrained peptides and peptide-polyketide hybrids, with a particular emphasis on those systems containing an ene-like bond.

There is an increasing demand for biocompatible materials in biomedical applications. Herein, we report a modified α-helical decapeptide segment from the cardiac troponin C, which self-assembles into fibers with a secondary β-sheet structure. These fibers cross-link via a novel supramolecular threading mechanism which results in an atypical stiff hydrogel (G' ≈ 13 kPa). In this work, we provide a first insight into the understanding of such remarkable cross-linking mechanism, which will aid in the development of new biomaterials with unique properties.

The first synthesis of the anti-TB cyclic peptide callyaerin A (1), containing a rare (Z)-2,3-diaminoacrylamide bridging motif, is reported. Fmoc-formylglycine-diethylacetal was used as a masked equivalent of formylglycine in the synthesis of the linear precursor to 1. Intramolecular cyclization between the formylglycine residue and the N-terminal amine in the linear peptide precursor afforded the macrocyclic natural product 1. Synthetic 1 possessed potent anti-TB activity (MIC100 =32 μm) while its all-amide congener was inactive. Variable-temperature NMR studies of both the natural product and its all-amide analogue revealed the extraordinary rigidity imposed by this diaminoacrylamide unit on peptide conformation. The work reported herein pinpoints the intrinsic role that the (Z)-2,3-diaminoacrylamide moiety confers on peptide bioactivity.