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Phytosulfokine (PSK) is a highly conserved plant peptide hormone (PPH) containing two sulfated tyrosine residues. PSK acts as a growth factor in Arabidopsis thaliana plant species and significantly improves the efficiency of cellular proliferation in plant cell culture, a process known as regeneration. However, many important plant species are recalcitrant under exposure to PSK and require e.g., artificially generated PPHs to boost regeneration. Here, we report the design, chemical synthesis, and biological evaluation of the regeneration capacity of a PSK-like peptide library. Our set of thirteen PSK analogues included N-terminal modifications, amino acid substitutions, incorporation of diastereomers, and backbone N-methylation. Insertion of the latter required solution-phase synthesis of the proper building block prior to solid-phase peptide synthesis. The biological activity of the PSK analogues was assessed in the regeneration capacity of Brassica oleracea cells (protoplasts). Most PSK analogues did not affect the regeneration capacity as compared to PSK. For example, when an alanine substitution was introduced and replaced either 2-Ile or 4-Thr, the bio-activity was fully abolished. However, 2-allo-Ile PSK and 4-Nme-Thr PSK showed increased activity in protoplast regeneration when compared with that of the native peptide. These findings indicated that introducing small chemical modifications in the PPH PSK provides opportunities for inducing protoplast regeneration in recalcitrant species. [ABSTRACT FROM AUTHOR]

Self-assembled peptide hydrogels possess remarkable biological activity and biocompatibility. When utilized for local drug delivery, they require enhanced mechanical properties and the ability to facilitate controlled release of drugs. Herein, we present the design and synthesis of a supramolecular hydrogel (Fmoc-GFFG/SBE-β-CD). The hydrogel incorporates host–guest interactions between a self-assembling peptide (Fmoc-GFFG) and cyclodextrin molecules (SBE-β-CD) to improve the viscoelastic strength of the hydrogel while maintaining biocompatibility. With remarkable shear-thinning properties, the Fmoc-GFFG/SBE-β-CD hydrogel exhibits enhanced in vivo retention time, thereby facilitating the development of an injectable drug delivery system. Using 10-hydroxycamptothecin (HCPT) as a model drug, we develop a hydrogel system (Fmoc-GFFG/SBE-β-CD@HCPT) that capitalizes on the inclusion complex formation between SBE-β-CD and HCPT. The resulting composite hydrogels effectively prevent the agglomeration of drug molecules, ensuring the sustained release of HCPT while preserving its anticancer efficacy. The findings presented in this work establish a foundation for the broadening of peptide-based biomaterials and offer a promising avenue for the development of sophisticated drug delivery systems. [ABSTRACT FROM AUTHOR]

Pseudo-allergic reactions are non-IgE-mediated immediate allergic reactions, and MRGPRX2 is a potential important target for pseudo-allergic reactions. Currently, numerous reports focus on the induction of pseudo-allergic reactions by small molecule drugs and endogenous peptides. However, insufficient attention has been paid to the pseudo-allergic reactions induced by exogenous peptides through MRGPRX2. Therefore, this study selected immunogenic peptide composition HL-9-5, which has a structure and properties similar to substance P, and its analogue HL-9-4. We conducted experiments through LAD2 cell β-hexosaminidase release, histamine release, cytokine release and MRGPRX2-HEK293 cell calcium mobilization assay in vitro, and mice paw swelling, serum histamine detection, hindpaw skin H&E and TB staining in vivo demonstrated that HL-9-4 and HL-9-5 can trigger pseudo-allergic reactions via MRGPRX2. Molecular docking further showed that SP, HL-9-4 and HL-9-5 can act on the same cavity in MRGPRX2, and ASP-174 was identified as the conserved site of interaction for all three compounds. Therefore, HL-9-4 and HL-9-5 are exogenous analogues of substance P that can induce pseudo-allergic reactions. This study laid the foundation for research on the mechanism of pseudo-allergic reactions induced by exogenous peptide compounds and was also of great significance for subsequent research on the interaction between various ligands and MRGPRX2 and for the development of peptide antagonists. [ABSTRACT FROM AUTHOR]

Peptides respresent intriguing materials to achieve sustainable catalytic reactivity that mimic the natural functions of enzymes, but without the limitations of temperature/solvent sensitivity. They could also be applicable to a wide variety of substrates, thus expanding their potential use at different reaction levels ranging from the benchtop to industrial. Unfortunately, significant use of catalytic peptides remains limited due to the general lack of understanding of the fundamental basis of their inherent reactivity. In this contribution, we examine the reactivity of a peptide (termed CPN3) previously isolated with ester hydrolysis reactivity. It is demonstrated that the system is most reactive under slightly basic conditions. While the system is slower than comparable enzymes, it demonstrates significant reactivity across multiple substrates and different reaction conditions that could likely lead to enzymatic denaturation. In addition, key active site residues were identified to begin to elucidate the fundamental basis of the reactivity. Such results could be used to design new sequences with enhanced reactivity under sustainable conditions. [ABSTRACT FROM AUTHOR]

Water soluble, cationic charge, peptide appended perylenebisimide (PBI-CFK) has been designed, synthesized, purified, and fully characterized. A hybrid system has been composed with PBI-CFK and oppositely charged deprotonated ferrocenedicarboxylic acid (Fc). In this case, PBI-CFK acts as an acceptor and Fc as a donor. The UV-visible study reveals the blue shift of PBI-CFK absorption and its photoluminescence intensity is gradually quenched upon the addition of Fc upto 1 equivalent. HR-TEM (high-resolution transmission electron microscopy) studies exhibit highly ordered and more condensed structures of the hybrid system than those of each of these components. The current–voltage (I–V) study clearly shows that the hybrid provides better semiconducting properties compared to PBI-CFK, which appears to be due to the electronic and ionic interaction of these two components. Here, PBI-CFK exhibits photoswitching properties with a photocurrent gain (Ion/Ioff) of 2.04, whereas the hybrid shows a significant improvement of the photoswitching properties with the photocurrent gain (Ion/Ioff) of 4.04 and this is about twice that of the individual one. Thus, the formation of a donor–acceptor conjugate between Fc and PBI-CFK results in modulation of the optoelectronic and photoswitching properties. [ABSTRACT FROM AUTHOR]

Recently, we have studied the coordination chemistry of the Cu(II)–histidine-rich C-terminal tail (HRCT) complex of the mycobacterial GroEL1 protein. The structure of this domain differs significantly compared to the well-known methionine-glycine-rich GroEL chaperonin – it was predicted that mycobacterial GroEL1 could play a significant role in the metal homeostasis of Mycobacteria, especially copper. However, we found that this particular domain's pattern also repeats in a number of Ni(II)-binding proteins. Here, we present the studies concerning the properties of GroEL1 HRCT as a ligand for Ni(II) ions. For this purpose, we chose eight model peptides: L1 – Ac-DHDHHHGHAH, L2 – Ac-DKPAKAEDHDHHHGHAH, and 6 mutants of the latter in the pH range of 2–11. We examined the stoichiometry, stability, and spectroscopic features of copper complexes. We noticed that similar to the Cu(II)-complex, the presence of a Lys5 residue significantly increases the stability of the system. The impact of His mutations was also examined and carefully studied using NMR spectroscopy. His9 and His13 are the crucial residues for Ni(II) binding, whereas His12 has minimal relevance in complex formation. [ABSTRACT FROM AUTHOR]

Hydrogen/deuterium exchange mass spectrometry (HDX-MS) is a versatile bioanalytical technique for protein analysis. Since the reliability of HDX-MS analysis considerably depends on the retention of deuterium labels in the post-labeling workflow, deuterium/hydrogen (D/H) back exchange prevention strategies, including decreasing the pH, temperature, and exposure time to protic sources of the deuterated samples, are widely adopted in the conventional HDX-MS protocol. Herein, an alternative and effective back exchange prevention strategy based on the encapsulation of a millimeter droplet of a labeled peptide solution in a water-immiscible organic solvent (cyclohexane) is proposed. Cyclohexane was used to prevent the undesirable uptake of water by the droplet from the atmospheric vapor through the air–water interface. Using the pepsin digest of deuterated myoglobin, our results show that back exchange kinetics of deuterated peptides is retarded in a millimeter droplet as compared to that in the bulk solution. Performing pepsin digestion directly in a water-in-oil droplet at room temperature (18–21 °C) was found to preserve more deuterium labels than that in the bulk digestion with an ice-water bath. Based on the present findings, it is proposed that keeping deuterated peptides in the form of water-in-oil droplets during the post-labelling workflow will facilitate the preservation of deuterium labels on the peptide backbone and thereby enhance the reliability of the H/D exchange data. [ABSTRACT FROM AUTHOR]

The detection of copper ions and hydrogen sulfide with high sensitivity and selectivity has become a research priority as they play a crucial role in physiopathological processes both in the human body and in the environment. Herein, a fluorescent chemosensor named FGP1 (FITC-Ahx-Ser-Ser-His-Thr-Glu-Phe-NH2) was developed using novel peptides. This chemosensor solely recognizes Cu2+ without any disruption by other metal ions that may compete. The binding of FGP1–Cu leads to the quenching of the FGP1 probe fluorescence with great sensitivity and rapid response time, resulting in a noticeable change of color observable to the naked eye. The association constant of FGP1–Cu is 7.34 × 105 M−1, and thus when S2− is added to the FGP1–Cu complexes, Cu2+ is removed and fluorescence is restored, resulting in an "off–on" process. The fluorescence titration measurements' detection limits, computed utilizing the 3σ/k equation, were 38.2 nM (Cu2+) and 37.9 nM (S2−). These values are considerably lower than the safe levels designated by the US EPA. Furthermore, on account of its favorable water solubility, high biocompatibility and low toxicity, the FGP1 probe proved effective in tracking Cu2+ and S2− in HeLa living cells using the fluorescence imaging technique. [ABSTRACT FROM AUTHOR]

Mycobacterial histidine-rich GroEL1 protein significantly differs from the well-known methionine-glycine-rich GroEL chaperonin and most preferably participates in Cu(II) homeostasis. Some GroEL1 proteins, however, do not possess six but only three histidine residues and more acidic residues that can function as binding sites for metal ions. To evaluate the importance of this difference, we examined and compared the properties of GroEL1 His-rich or Glu/His-rich C-terminal domains as ligands for Cu(II), Ni(II), and Zn(II) ions. We studied the stoichiometry, stability, and binding sites of Cu(II)/Ni(II)/Zn(II) complexes of two model peptides: XEN = Ac-DKPEEEEDGHGHAH (M. xenopi) and ABS = Ac-DKPAEEADHGHGHHGHAH (M. abscessus) in the pH range 2–11. In the case of Cu(II), Ni(II), and Zn(II) complexes of XEN and ABS, ABS always formed more stable complexes. For XEN, there seemed to be no preference for Ni(II) or Zn(II) ions. In contrast, for ABS, Zn(II) formed a complex that was slightly more stable than the one formed by Ni(II). This may be due to the 6 His residues, which preferentially interact with Zn(II) rather than Ni(II). The study identified that an equilibrium of complexes—known as polymorphism—may occur in ABS complexes. Therefore, distinct sets of histidine residues may be involved in metal binding. [ABSTRACT FROM AUTHOR]

Protein S-acylation is an important lipid modification characteristic for heterogeneity in the acyl chain and dynamicity in the acylation/deacylation cycle. Most S-acylproteomic research has been limited by indirect identification of modified proteins/peptides without attached fatty acids, resulting in the failure to precisely characterize S-acylated sites with attached fatty acids. The study of S-acylation turnover is still limited at the protein level. Herein, aiming to site-specifically profile both the heterogeneity and the turnover of S-acylation, we first developed a site-specific strategy for intact S-acylated peptide analysis by introducing an acid cleavable bioorthogonal tag into a metabolic labelling method (ssMLCC). The cleavable bioorthogonal tag allowed for the selective enrichment and efficient MS analysis of intact S-acylated peptides so that S-acylated sites and their attached fatty acids could be directly analysed, enabling the precise mapping of S-acylated sites, as well as circumventing false positives from previous studies. Moreover, 606 S-palmitoylated (C16:0) sites of 441 proteins in HeLa cells were identified. All types of S-acylated peptides were further characterized by an open search, providing site-specific profiling of acyl chain heterogeneity, including S-myristoylation, S-palmitoylation, S-palmitoleylation, and S-oleylation. Furthermore, site-specific monitoring of S-palmitoylation turnover was achieved by coupling with pulse-chase methods, facilitating the detailed observation of the dynamic event at each site in multi-palmitoylated proteins, and 85 rapidly cycling palmitoylated sites in 79 proteins were identified. This study provided a strategy for the precise and comprehensive analysis of protein S-acylation based on intact S-acylated peptide analysis, contributing to the further understanding of its complexity and biological functions. [ABSTRACT FROM AUTHOR]

Antimicrobial peptides show promise in combating microbial infections in the age of rapidly growing resistance against antibiotics. The salt sensitivity of their activity, protease susceptibility, and cytotoxicity are some of the disadvantages that limit the commercial success of AMPs. In this study, the hydrophobic–hydrophilic balance of lipopeptides was tuned to generate a highly potent AMP, P18, which overcomes the limitations of the existing AMPs. P18 is a non-cytotoxic, salt-tolerant, and protease-resistant AMP, with no development of resistance over 16 generations in MRSA, which is highly potent against ESKAPE pathogens such as P. aeruginosa, K. pneumoniae, S. aureus, and methicillin-resistant S. aureus (MRSA). This study explores and establishes the direct effect of the hydrophobic–hydrophilic balance of peptides on their various properties such as secondary structure, self-assembly (morphology and kinetics), antimicrobial potency (salt-tolerance, kinetics, etc.), and protease-resistance ability. Thus, peptides with tailor-made properties and functions can be designed in the future by being able to control and tune their overall hydrophobic–hydrophilic balance. [ABSTRACT FROM AUTHOR]

Superoxide dismutase (SOD) enzymes are pivotal in regulating oxidative stress. In order to model Ni containing SOD enzymes, the results of the thermodynamic, spectroscopic and SOD activity studies on the complexes formed between nickel(II) and a NiSOD related peptide, CysCysAspLeuProCysGlyValTyr–NH2 (wtCC), are reported. Cysteine was introduced to replace the first histidine residue in the amino acid sequence of the active site of the NiSOD enzyme. The novel peptide exhibits 3 times higher metal binding affinity compared to the native NiSOD fragment. This is due to the presence of the first cysteine in the coordination sphere of nickel(II). At physiological pH, the (NH2,S–,S–,S–) coordinated complex is the major species. This coordination mode is altered when one thiolate group is replaced by an amide nitrogen of the peptide backbone above pH 7.5. The nickel complexes of wtCC exhibit similar SOD activity to that of the complex formed with the active site fragment of the native NiSOD. The reaction between the complexes and the superoxide anion was studied by the sequential stopped-flow method. These studies revealed that the nickel(II) complex is always in excess over the nickel(III) complex during the dismutation process. However, the nickel(III) species is also involved in a relatively fast degradation process. This unambiguously proves that a protective mechanism must be operative in the NiSOD enzyme which prevents the oxidation of the sulfur atom of cysteine in the presence of O2−. The results provide new possibilities for the use of NiSOD mimics in bio- and industrial catalytic processes. [ABSTRACT FROM AUTHOR]

Caspase-3 is an important biomarker for the process of apoptosis, which is a key target for cancer treatment. Due to its low concentration in single cells and the structural similarity of caspase family proteins, it is exceedingly challenging to accurately determine the intracellular caspase-3 during apoptosis in situ. Herein, a biosensing strategy based on the target-induced SERS "hot spot" formation has been developed for the simultaneous highly sensitive and selective detection of intracellular caspase-3 level. The nanosensor is composed of gold nanoparticles modified with the probe molecule 4-mercaptophenylboronic acid (4-MPBA) and a peptide chain. The well-designed peptide chain contains two distinct functional domains, one with a sulfhydryl group for bonding to the gold nanoparticles and the other a fragment specifically recognized by caspase-3. When caspase-3 is present, the negatively charged segment (NH2–Asp–Asp–Asp–Glu–Val–Asp–OH) of the peptide chain is specifically hydrolyzed, leaving a positively charged fragment coated on the surface of the gold nanoparticles. At this time, the golden nanoparticles undergo significant coupling aggregation due to the electrostatic interaction, resulting in a large number of SERS "hot spot" formation. The SERS signal of the 4-MPBA located at the nano-gap is significantly boosted because of the local plasma enhancement effect. The highly sensitive determination of caspase-3 can be achieved according to the altered SERS signal intensity of 4-MPBA. The turn-on of the SERS signal-induced target contributes to the excellent selectivity and the formation of the SERS "hot spot" effect that further improves the sensitivity of caspase-3 detection. The advantages of this biosensing technique allow for the precise in situ monitoring of the dynamic changes in caspase-3 levels during apoptosis. In addition, the differences in caspase-3 levels during the apoptosis of various cell types were compared. Monitoring the caspase-3 levels can be used to track the cellular apoptosis process, evaluate the effect of drugs on cancer cells in real time, and provide guidance for the selection of the appropriate drug dosage. [ABSTRACT FROM AUTHOR]

Covering: 1992 up to 2023 Since their discovery, lasso peptides went from peculiarities to be recognized as a major family of ribosomally synthesized and post-translationally modified peptide (RiPP) natural products that were shown to be spread throughout the bacterial kingdom. Microcin J25 was first described in 1992, making it one of the earliest known lasso peptides. No other lasso peptide has since then been studied to such an extent as microcin J25, yet, previous review articles merely skimmed over all the research done on this exceptional lasso peptide. Therefore, to commemorate the 30th anniversary of its first report, we give a comprehensive overview of all literature related to microcin J25. This review article spans the early work towards the discovery of microcin J25, its biosynthetic gene cluster, and the elucidation of its three-dimensional, threaded lasso structure. Furthermore, the current knowledge about the biosynthesis of microcin J25 and lasso peptides in general is summarized and a detailed overview is given on the biological activities associated with microcin J25, including means of self-immunity, uptake into target bacteria, inhibition of the Gram-negative RNA polymerase, and the effects of microcin J25 on mitochondria. The in vitro and in vivo models used to study the potential utility of microcin J25 in a (veterinary) medicine context are discussed and the efforts that went into employing the microcin J25 scaffold in bioengineering contexts are summed up. [ABSTRACT FROM AUTHOR]

Self-assembly is an important strategy for constructing ordered structures and complex functions in nature. Based on this, people can imitate nature and artificially construct functional materials with novel structures through the supermolecular self-assembly pathway of biological interfaces. Among the many assembly units, peptide molecular self-assembly has received widespread attention in recent years. In this review, we introduce the interactions (hydrophobic interaction, hydrogen bond, and electrostatic interaction) between peptide nanomaterials and biological interfaces, summarizing the latest advancements in multifunctional self-assembling peptide materials. We systematically demonstrate the assembly mechanisms of peptides at biological interfaces, such as proteins and cell membranes, while highlighting their application potential and challenges in fields like drug delivery, antibacterial strategies, and cancer therapy. [ABSTRACT FROM AUTHOR]

Existing therapies for neurodegenerative diseases like Parkinson's and Alzheimer's address only their symptoms and do not prevent disease onset. Common therapeutic agents, such as small molecules and antibodies struggle with insufficient selectivity, stability and bioavailability, leading to poor performance in clinical trials. Peptide-based therapeutics are emerging as promising candidates, with successful applications for cardiovascular diseases and cancers due to their high bioavailability, good efficacy and specificity. In particular, cyclic peptides have a long in vivo stability, while maintaining a robust antibody-like binding affinity. However, the de novo design of cyclic peptides is challenging due to the lack of long-lived druggable pockets of the target polypeptide, absence of exhaustive conformational distributions of the target and/or the binder, unknown binding site, methodological limitations, associated constraints (failed trials, time, money) and the vast combinatorial sequence space. Hence, efficient alignment and cooperation between disciplines, and synergies between experiments and simulations complemented by popular techniques like machine-learning can significantly speed up the therapeutic cyclic-peptide development for neurodegenerative diseases. We review the latest advancements in cyclic peptide design against amyloidogenic targets from a computational perspective in light of recent advancements and potential of machine learning to optimize the design process. We discuss the difficulties encountered when designing novel peptide-based inhibitors and we propose new strategies incorporating experiments, simulations and machine learning to design cyclic peptides to inhibit the toxic propagation of amyloidogenic polypeptides. Importantly, these strategies extend beyond the mere design of cyclic peptides and serve as template for the de novo generation of (bio)materials with programmable properties. [ABSTRACT FROM AUTHOR]