Stable Peptide Research Articles

Chemical strategies to tune potency, selectivity and stability of venom peptides

Chemical strategies to tune potency, selectivity and stability of venom peptides

Origami of KR-12 Designed Antimicrobial Peptides and Their Potential Applications.

This review describes the discovery, structure, activity, engineered constructs, and applications of KR-12, the smallest antibacterial peptide of human cathelicidin LL-37, the production of which can be induced under sunlight or by vitamin D. It is a moonlighting peptide that shows both antimicrobial and immune-regulatory effects. Compared to LL-37, KR-12 is extremely appealing due to its small size, lack of toxicity, and narrow-spectrum antimicrobial activity. Consequently, various KR-12 peptides have been engineered to tune peptide activity and stability via amino acid substitution, end capping, hybridization, conjugation, sidechain stapling, and backbone macrocyclization. We also mention recently discovered peptides KR-8 and RIK-10 that are shorter than KR-12. Nano-formulation provides an avenue to targeted delivery, controlled release, and increased bioavailability. In addition, KR-12 has been covalently immobilized on biomaterials/medical implants to prevent biofilm formation. These constructs with enhanced potency and stability are demonstrated to eradicate drug-resistant pathogens, disrupt preformed biofilms, neutralize endotoxins, and regulate host immune responses. Also highlighted are the safety and efficacy of these peptides in various topical and systemic animal models. Finaly, we summarize the achievements and discuss future developments of KR-12 peptides as cosmetic preservatives, novel antibiotics, anti-inflammatory peptides, and microbiota-restoring agents.

Cellular N-Myristoyl Transferases Are Required for Mammarenavirus Multiplication.

The mammarenavirus matrix Z protein plays critical roles in virus assembly and cell egress. Meanwhile, heterotrimer complexes of a stable signal peptide (SSP) together with glycoprotein subunits GP1 and GP2, generated via co-and post-translational processing of the surface glycoprotein precursor GPC, form the spikes that decorate the virion surface and mediate virus cell entry via receptor-mediated endocytosis. The Z protein and the SSP undergo N-terminal myristoylation by host cell N-myristoyltransferases (NMT1 and NMT2), and G2A mutations that prevent myristoylation of Z or SSP have been shown to affect the Z-mediated virus budding and GP2-mediated fusion activity that is required to complete the virus cell entry process. In the present work, we present evidence that the validated on-target specific pan-NMT inhibitor DDD85646 exerts a potent antiviral activity against the prototypic mammarenavirus lymphocytic choriomeningitis virus (LCMV) that correlates with reduced Z budding activity and GP2-mediated fusion activity as well as with proteasome-mediated degradation of the Z protein. The potent anti-mammarenaviral activity of DDD85646 was also observed with the hemorrhagic-fever-causing Junin (JUNV) and Lassa (LASV) mammarenaviruses. Our results support the exploration of NMT inhibition as a broad-spectrum antiviral against human pathogenic mammarenaviruses.

Amine-bearing hydrocarbon cross-links: Tailoring helix stability, hydrophilicity, and synthetic adaptability in peptides

This study comprehensively explored the helix-stabilizing effects of amine-bearing hydrocarbon cross-links (ABXs), revealing their context-dependent nature influenced by various structural parameters. Notably, we identified a 9-atom ABX as a robust helix stabilizer, showcasing versatile synthetic adaptability while preserving peptide water solubility. Future investigations are imperative to fully exploit this system’s potential and enrich our chemical toolkit for designing innovative peptide-based biomolecules.

A Critical Review on the Identification of Pathogens by Employing Peptide-Based Electrochemical Biosensors.

In this era of emerging pathogenic diseases, prompt and accurate detection of pathogens is crucial. Disease diagnosis, environmental monitoring and food safety all rely heavily on the identification of pathogens. Peptide-based electrochemical sensors due to their rapid response times, specificity and sensitivity have emerged as promising tools in the identification of pathogens. This review emphasizes the importance of peptides in detection of pathogens and different peptide-based electrochemical biosensors for the detection of pathogens. Peptides offer several advantages including strong binding affinity to a diverse array of pathogens including bacteria, viruses and fungi, tunable specificity and simple synthesis. Peptide-based electrochemical sensors employ different electrochemical techniques such as differential pulse voltammetry (DPV), cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS), amperometry and linear sweep voltammetry (LSV). The efficacy of peptide-based biosensors in detecting low concentrations of pathogens is highlighted, demonstrating the promising applications of these biosensors in early diagnosis and real-time monitoring. In addition, the review also addresses the current challenges in the field such as peptide stability, sensor reproducibility and interference from complex biological matrices. This review suggests potential resolutions and avenues for progress such as the development of multiplexed detection systems that can concurrently identify multiple pathogens and developments in peptide design and sensor miniaturization. In summary, this review highlights the substantial advancements and potential possibilities of peptide-based electrochemical biosensors in the realm of pathogen detection, thereby facilitating the development of safer and more effective diagnostic tools.

A computational approach to identifying peptide inhibitors againstWhite Spot Syndrome Virus: Targeting the virus envelope protein

The white spot syndrome virus (WSSV), a rapidly replicating and highly lethal pathogen that targets Penaeid shrimp, has emerged as one of the most widespread viruses globally due to its high virulence. With effective chemotherapeutics still unavailable, the pursuit of novel and viable strategies against WSSV remains a crucial focus in the field of shrimp farming. The envelope proteins of WSSV are essential for virus entry, serving as excellent targets for the development of antiviral therapeutics. Novel strategies in the design of inhibitory peptides, especially those targeting envelope protein (VP28) located on the surface of the virus particle, play a critical role as a significant virulence factor during the early stages of inherent WSSV infection in shrimp. In this direction, the current computational study focused on identifying self-inhibitory peptides from the hydrophobic membrane regions of the VP28 protein, employing peptide docking and molecular dynamics simulation (MDS) approaches. Such inhibitory peptides could be useful building blocks for the rational engineering of inhibitory therapeutics since they imitate the mechanism of binding to homologous partners used by their origin domain to interact with other molecules. The N-terminal sequence of VP28 has been reported as the potential site for membrane interactions during the virus entry. Moreover, drug delivery systems mediated by chitosan and gold nanoparticles are being developed to enhance the therapeutic efficacy of anti-viral peptides. These systems can increase the solubility, stability, and selectivity of peptides, possessing better qualities than conventional delivery methods. This computational study on self-inhibitory peptides could be a valuable resource for further in vitro and in vivo studies on anti-viral therapeutics in the aquaculture industry.

PrecivityAD2™ Blood Test: Analytical Validation of an LC-MS/MS Assay for Quantifying Plasma Phospho-tau217 and Non-Phospho-tau217 Peptide Concentrations That Are Used with Plasma Amyloid-β42/40 in a Multianalyte Assay with Algorithmic Analysis for Detecting Brain Amyloid Pathology.

Alzheimer's disease (AD) is a progressive irreversible neurodegenerative disorder that represents a major global public health concern. Traditionally, AD is diagnosed using cerebrospinal fluid biomarker analysis or brain imaging modalities. Recently, less burdensome, more widely available blood biomarker (BBM) assays for amyloid-beta (Aβ42/40) and phosphorylated-tau concentrations have been found to accurately identify the presence/absence of brain amyloid plaques and tau tangles and have helped to streamline AD diagnosis. However, few BBMs have been rigorously analytically validated. Herein, we report the analytical validation of a novel liquid chromatography-tandem mass spectrometry (LC-MS/MS) multiplex method for quantifying plasma phosphorylated-tau217 (p-tau217) and non-phosphorylated-tau217 (np-tau217) peptide concentrations. We combined the p-tau217/np-tau217 concentrations ratio (%p-tau217) and the previously validated LC-MS/MS multiplex assay for plasma Aβ42/40 into a new multianalyte assay with algorithmic analysis (MAAA; PrecivityAD2™ test) that identifies brain amyloid status based on brain amyloid positron emission tomography. We found (a) the %p-tau217 assay is precise, accurate, sensitive, and linear over a wide analytical measurement range, and free from carryover and interference; (b) the pre-analytical specimen collection, processing, storage, and shipping conditions that maintain plasma tau peptide stability; and (c) using the measured analytical imprecision for plasma Aβ42/40 and p-tau217/np-tau217 levels in a worst-case scenario model, the PrecivityAD2 test algorithm for amyloid pathology classification changed for only 3.5% of participants from brain amyloid positive to negative, or from negative to positive. The plasma sample preparation and LC-MS/MS methods underlying the PrecivityAD2 test are suitable for use in the clinical laboratory and valid for the test's intended purpose: to aid in the diagnostic evaluation of individuals aged 55 and older with signs or symptoms of mild cognitive impairment or dementia.

From self-Assembly to healing: Engineering ultra-Small peptides into supramolecular hydrogels for controlled drug release

The aim of this study was the evaluation of suitability of novel mucoadhesive hydrogel platforms for the delivery of therapeutics useful for the management of disorders related to the gastrointestinal tract (GI). At this purpose, here we describe the preparation, the physicochemical characterization and drug delivery behaviour of novel hydrogels, based on self-assembling lipopeptides (MPD02-09), obtained by covalently conjugating lauric acid (LA) to SNA’s peptide derivatives gotten by variously combining D- and L- amino acid residues. LA conjugation was aimed at improving the stability of the precursor peptides, obtaining amphiphilic structures, and triggering the hydrogels formation through the self-assembling. Budesonide (BUD), an anti-inflammatory drug, was selected as model because of its use in the treatment in GI disorders. Preliminary studies were performed to correlate the chemical structure of the conjugates with the key physicochemical properties of the materials for drug delivery. Two lipopeptides, MPD03 and MPD08, were found to form hydrogels (MPD03h and MPD08h, respectively) with characteristics suitable for drug delivery. These materials showed mucoadhesiveness of about 60 %. In vitro studies carried out with BUD loaded hydrogels showed about 70 % drug release within 6 h. Wound healing assessed in Caco-2 and HaCaT cells, showed reduction of cell-free area to values lower than 10 %. Taking together these results MPD03h and MPD08h have been shown to be excellent candidates for BUD delivery.

Abstract Tu005: Small Extracellular Vesicles Derived from Bovine Milk Contain an Endogenous Carboxyl Terminal Polypeptide of The Gap Junction Protein Connexin-43

Background: Connexin 43 (Cx43) is the body's most widely expressed gap junctional connexin. Researchers have shown evidence of non-canonical functions of Cx43 and the therapeutic promise of Cx43 mimetic peptides in cardiac pathologies. However, some critical barriers, including in vivo stability and potential immunotoxicity of Cx43 mimetic peptides, hinder clinical translation of these pro-drugs. To address these challenges, we are pioneering bovine milk-derived small extracellular vesicles (mEVs) as delivery vehicles for peptidic drugs. While exploring mEVs as drug delivery tools for αCT1 – a peptide mimicking Cx43 carboxyl terminus (CT) undergoing Phase III testing and known for promoting wound healing and cardioprotection - mEVs without loaded drug cargo exhibited similar bioactivity. This study aimed to test the hypothesis that Cx43, particularly CT fragments of Cx43, are naturally present in mEVs. Methods: mEVs were isolated from bovine milk using a novel protocol developed by our lab based on Tangential Flow Filtration and characterized by Western Blotting (WB), Nanoparticle Tracking Analysis, and Transmission Electron Microscopy. To determine the presence of Cx43 in mEVs, we used WB, immunoprecipitation and Single-Molecule Localization Microscopy (SMLM). Results: Characterization of mEVs confirmed relatively pure and ultrastructurally definitive mEVs isolated from milk at high density. WB for Cx43 in mEVs lysates based on two antibodies - one against the cytoplasmic loop domain and the other against CT-most 20 amino acids of Cx43, showed that full-length Cx43 was present in mEVs. Interestingly, the Cx43 CT antibody also detected a band at around 16 KDa, and this was confirmed by WB by using two more Cx43 CT antibodies. We also immunoprecipitated this Cx43 CT polypeptide from mEVs lysates using the CT antibodies. SMLM suggested that a subpopulation of mEVs contain Cx43 and demonstrated vesicle-to-vesicle heterogeneity to Cx43 immunolabeling. Conclusions: We show for the first time that an endogenous Cx43 CT polypeptide is present in mEVs. Our results provide new insight into the role of Cx43 in mEV bioactivity and potentially offer a path to safe, effective, and orally available therapeutic options based on mEVs.

Cross-Species Applications of Peptide Substrate Reporters to Quantitative Measurements of Kinase Activity.

Peptide substrate reporters are short chains of amino acids designed to act as substrates for enzymes of interest. Combined with capillary electrophoresis and laser-induced fluorescence detection (CE-LIF), they are powerful molecular tools for quantitative measurements of enzyme activity even at the level of single cells. Although most peptide substrate reporters have been optimized for human or murine cells in health-related applications, their performance in nonmammalian organisms remains largely unexplored. In this study, we evaluated three peptide substrate reporters for protein kinase B (PKB) in two eukaryotic microbes, Dictyostelium discoideum and Tetrahymena thermophila, which are evolutionarily distant from mammals and from each other yet express PKB homologues. All three peptide substrate reporters were phosphorylated in lysates from both organisms but with varying phosphorylation kinetics and stability. To demonstrate reporter utility, we used one to screen for and identify the previously unknown stimulus needed to activate PHK5, the PKB homologue in T. thermophila. In D. discoideum, we employed the highly quantitative nature of these assays using CE-LIF to make precise measurements of PKB activity in response to transient stimulation, drug treatment, and genetic mutation. These results underscore the broad applicability of peptide substrate reporters across diverse species while highlighting the need for further research to determine effective peptide stabilization strategies across different biological contexts.

Cellular N-myristoyl transferases Are Required for Mammarenavirus Multiplication.

The mammarenavirus matrix Z protein plays critical roles in virus assembly and cell egress, whereas heterotrimer complexes of a stable signal peptide (SSP) together with glycoprotein subunits GP1 and GP2, generated via co-and post-translational processing of the surface glycoprotein precursor GPC, form the spikes that decorate the virion surface and mediate virus cell entry via receptor-mediated endocytosis. The Z protein and SSP undergo N-terminal myristoylation by host cell N-myristoyltransferases (NMT1 and NMT2), and G2A mutations that prevent myristoylation of Z or SSP have been shown to affect Z mediated virus budding and GP2 mediated fusion activity required to complete the virus cell entry process. In the present work, we present evidence that the validated on-target specific pan NMT inhibitor DDD85464 exerts a potent antiviral activity against the prototypic mammarenavirus lymphocytic choriomeningitis virus (LCMV) that correlated with reduced Z budding activity and GP2 mediated fusion activity, as well as proteasome mediated degradation of the Z protein. The potent anti-mammarenaviral activity of DDD85646 was also observed with the hemorrhagic fever causing mammarenaviruses Junin (JUNV) and Lassa (LASV) viruses. Our results support exploration of NMT inhibition as a broad-spectrum antiviral against human pathogenic mammarenaviruses.

Identification, screening and molecular mechanisms of natural stable angiotensin-converting enzyme (ACE) inhibitory peptides from foxtail millet protein hydrolysates: a combined in silico and in vitro study.

The stability of bioactive peptides under various food processing conditions is the basis for their use in industrial manufacturing. This study aimed to identify natural ACE inhibitors with excellent stability and investigate their physicochemical properties and putative molecular mechanisms. Five novel ACE inhibitory peptides (QDPLFPL, FPGVSPF, SPAQLLPF, LVPYRP, and WYWPQ) were isolated and identified using RP-HPLC and Nano LC-MS/MS with foxtail millet protein hydrolysates as the raw material. These peptides are non-toxic and exhibit strong ACE inhibitory activity in vitro (IC50 values between 0.13 mg mL-1 and 0.56 mg mL-1). In addition to QDPLFPL, FPGVSPF, SPAQLLPF, LVPYRP, and WYWPQ have excellent human intestinal absorption. Compared to FPGVSPF and SPAQLLPF, the stable helical structure of LVPYRP and WYWPQ allows them to maintain high stability under conditions that mimic gastrointestinal digestion and various food processing (temperatures, pH, sucrose, NaCl, citric acid, sodium benzoate, Cu2+, Zn2+, K+, Mg2+, Ca2+). The results of molecular docking and molecular dynamics simulation suggest that LVPYRP has greater stability and binding capacity to ACE than WYWPQ. LVPYRP might attach to the active pockets (S1, S2, and S1') of ACE via hydrogen bonds and hydrophobic interactions, then compete with Zn2+ in ACE to demonstrate its ACE inhibitory activity. The binding of LVPYRP to ACE enhances the rearrangement of ACE's active structural domains, with electrostatic and polar solvation energy contributing the most energy to the binding. Our findings suggested that LVPYRP derived from foxtail millet protein hydrolysates has the potential to be incorporated into functional foods to provide antihypertensive benefits.

Preparation, Characterization and Stability of Calcium-Binding Peptides Derived from Chicken Blood.

Calcium-binding peptides have gained significant attention due to their potential applications in various fields. In this study, we aimed to prepare, characterize, and evaluate the stability of calcium-binding peptides derived from chicken blood. Chicken hemoglobin peptides (CPs) were obtained by protease hydrolysis and were applied to prepare chicken hemoglobin peptide-calcium chelate (CP-Ca). The preparation conditions were optimized, and the characteristics and stability of CP-Ca were analyzed. The optimal chelating conditions were determined by single-factor and response surface tests, and the maximum calcium ion chelating rate was 77.54%. Amino acid analysis indicated that glutamic acid and aspartic acid motifs played an important role in the chelation of the calcium ions and CP. According to the characterization analysis, CP-Ca was a different substance compared with CP; calcium ions chelated CPs via the sites of carbonyl oxygen, carboxyl oxygen, and amino nitrogen groups; and after the chelation, the structure changed from a smooth homogeneous plate to compact granular. The stability analysis showed that CP-Ca was stable at different temperatures, pH, and gastrointestinal conditions. The study indicates that chicken blood is a promising source of peptide-calcium chelates, providing a theoretical basis for application in functional foods and improving the utilization value of chicken blood.

Design of Hepatitis C Protease Inhibitors Based on Knottin Fold

AbstractKnottins are very stable peptides (also called miniproteins) containing approximately 30 amino acid residues. Their structures are stabilized by three disulfide bonds forming a characteristic ‘pseudo‐knotted’ structure. This research was devoted to the design of potential inhibitors of human hepatitis C (HCV) NS3 protease by means of knottin fold using loop grafting, amino acid residues replacements and L/D amino acid epimerization. The initial structure used for design was a trypsin inhibitor from Momordica cochinchinensis seeds, MCOTI‐II. Modifications of the knottin template for optimization of the NS3 protease‐peptide interaction included deletion of several residues from the N‐terminus, replacement of residues in‐ and outside the inhibitor loop and replacement of certain L‐amino acids by their D‐stereoisomers. Stability of modified knottins and their complexes with HCV protease were evaluated by molecular dynamics simulation. Binding energy values for protein‐knottin complexes were calculated by MM‐GBSA method. The designed modified knottin characterized by the highest stability of the complex with HCV NS3 protease, was proposed for further experimental testing.

A Bottom-Up Liquid Chromatography-Tandem Mass Spectrometry Method for Therapeutic Drug Monitoring of Infliximab: Method Development, Comparison With 2 Enzyme-Linked Immunosorbent Assay Methods, and Evaluation of Anti-Drug Antibody Interference.

Therapeutic drug monitoring is recommended to optimize infliximab use and improve outcome in chronic inflammatory disorders. To describe a simple and affordable liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to measure infliximab in serum. Infliximab was measured using winged stable isotope-labeled peptides as internal standards. Linearity, lower limit of measuring interval, limit of detection, precision, accuracy, carryover, and ion suppression were evaluated. Method comparison against 2 enzyme-linked immunosorbent assay (ELISA) methods (Remsima Monitor and IDKmonitor Infliximab) and anti-drug antibody (ADA) interference were evaluated using clinical specimens from inflammatory bowel disease patients (N = 237). Analytical run time and sample preparation time were 5 minutes per sample and 3 hours per batch, respectively. Analytical measurement interval and limit of detection were 0.50 to 50.0 μg/mL (R2 = 0.998) and 0.25 μg/mL, respectively. The intraday and interday imprecision percentage coefficients of variation were less than 6.1%. Accuracy was 94.2% to 98.7%. No significant ion suppression or carryover was observed. Infliximab concentrations measured by LC-MS/MS showed good agreement with those measured by Remsima Monitor (mean percentage difference, 5.7%; 95% CI, -1.2% to 12.6%) but were markedly lower than those measured by IDKmonitor (-32.6%; -35.8% to -29.4%), demonstrating significant bias between ELISAs. Although a good agreement between LC-MS/MS and ELISA was observed for ADA-negative samples (-3.5%; -12.8% to 5.9%), a significant bias was observed for ADA-positive samples (13.6%; 1.7% to 25.6%). This simple, fast, and affordable LC-MS/MS method for infliximab quantitation could improve standardization of infliximab quantitation and optimization of infliximab use in patients with high-titer ADA.

Site-selective peptide bond hydrolysis and ligation in water by short peptide-based assemblies

The evolution of complex chemical inventory from Darwin's nutrient-rich warm pond necessitated rudimentary yet efficient catalytic folds. Short peptides and their self-organized microstructures, ranging from spherical colloids to amyloidogenic aggregates might have played a crucial role in the emergence of contemporary catalytic entities. However, the question of how short peptide fragments had functions akin to contemporary complex enzymes to catalyze cleavage and formation of highly stable peptide bonds that constitute the backbone of all proteins remains an unresolved yet fundamentally important question in terms of the origins of enzymes. We report short-peptide-based spherical assemblies that demonstrated residue-specific cleavage and formation of peptide bonds of diverse peptide-based substrates under aqueous environment. Despite the short sequence length, the assemblies utilized the synergistic collaboration of four residues which included the catalytic triad of extant serine proteases with a nonproteinogenic amino acid (quinone moiety), to facilitate proteolysis, ligation, and a three-step (hydrolysis-ligation-hydrolysis) cascade. Such short-peptide-based catalytic assemblies argue for their candidacy as the earliest protein folds and open up avenues for biotechnological applications.

Cyclic tachyplesin I kills proliferative, non-proliferative and drug-resistant melanoma cells without inducing resistance

Acquired drug resistance is the major cause for disease recurrence in cancer patients, and this is particularly true for patients with metastatic melanoma that carry a BRAF V600E mutation. To address this problem, we investigated cyclic membrane-active peptides as an alternative therapeutic modality to kill drug-tolerant and resistant melanoma cells to avoid acquired drug resistance. We selected two stable cyclic peptides (cTI and cGm), previously shown to have anti-melanoma properties, and compared them with dabrafenib, a drug used to treat cancer patients with the BRAF V600E mutation. The peptides act via a fast membrane-permeabilizing mechanism and kill metastatic melanoma cells that are sensitive, tolerant, or resistant to dabrafenib. Melanoma cells do not become resistant to long-term treatment with cTI, nor do they evolve their lipid membrane composition, as measured by lipidomic and proteomic studies. In vivo studies in mice demonstrated that the combination treatment of cTI and dabrafenib resulted in fewer metastases and improved overall survival. Such cyclic membrane-active peptides are thus well suited as templates to design new anticancer therapeutic strategies.

Molecular peptide grafting as a tool for creating new generation of biopeptides: A mini-review

Molecular peptide grafting (MPG) is the isolation/synthesis of a bioactive fragment of a peptide/protein and its subsequent transfer to a target protein/peptide to create a new protein product with specified unique biological properties. This is one of the methods together with molecular stapling and peptide backbone circularization to strengthen the structural organization of short peptides. Nowadays research on MPT is mainly focused on demonstrating its usefulness and applicability, rather than on the development of next-generation biopeptides. The purpose of the mini-review is to demonstrate the applicability of MPT to create stable and bioavailable peptides of a new generation with enhanced biological properties. Choosing the right scaffold for subsequent inoculation of a biologically active peptide sequence into it is the most important task in creating targeted biopeptides. Peptides with the necessary framework, such as cyclotides, can be obtained by three-phase synthesis. Cyclotides have a common mechanism of action. Their biological activity is determined both by the ability to bind proteins with the formation of pores and destruction of biological target-membranes, and by the properties necessary to create new peptides in the scaffold. Various peptide inserts can be used to ensure the functionality of new biopeptides obtained by the MPT method. Different peptide drugs are an example of the effective practical use of MTP. Consequently, MPT makes it possible to effectively design a new generation of biopeptides characterized by high epitope thermodynamic and metabolic stability with new or enhanced biological functions. However, the effectiveness of the peptides obtained by the MPT must be proved in vitro and in vivo.

Differential potassium channel inhibitory activities of a novel thermostable degradation peptide BmKcug1a-P1 from scorpion medicinal material and its N-terminal truncated/restored peptides

Thermally stable full-length scorpion toxin peptides and partially degraded peptides with complete disulfide bond pairing are valuable natural peptide resources in traditional Chinese scorpion medicinal material. However, their pharmacological activities are largely unknown. This study discovered BmKcug1a-P1, a novel N-terminal degraded peptide, in this medicinal material. BmKcug1a-P1 inhibited hKv1.2 and hKv1.3 potassium channels with IC50 values of 2.12 ± 0.27 μM and 1.54 ± 0.28 μM, respectively. To investigate the influence of N-terminal amino acid loss on the potassium channel inhibiting activities, three analogs (i.e., full-length BmKcug1a, BmKcug1a-P1-D2 and BmKcug1a-P1-D4) of BmKcug1a-P1 were prepared, and their potassium channel inhibiting activities on hKv1.3 channel were verified by whole-cell patch clamp technique. Interestingly, the potassium channel inhibiting activity of full-length BmKcug1a on the hKv1.3 channel was significantly improved compared to its N-terminal degraded form (BmKcug1a-P1), while the activities of two truncated analogs (i.e., BmKcug1a-P1-D2 and BmKcug1a-P1-D4) were similar to that of BmKcug1a-P1. Extensive alanine-scanning experiments identified the bonding interface (including two key functional residues, Asn30 and Arg34) of BmKcug1a-P1. Structural and functional dissection further elucidated whether N-terminal residues of the peptide are located at the bonding interface is important in determining whether the N-terminus significantly influences the potassium channel inhibiting activity of the peptide. Altogether, this research identified a novel N-terminal degraded active peptide, BmKcug1a-P1, from traditional Chinese scorpion medicinal material and elucidated how the N-terminus of peptides influences their potassium channel inhibiting activity, contributing to the functional identification and molecular truncation optimization of full-length and degraded peptides from traditional Chinese scorpion medicinal material Buthus martensii Karsch.

Exploring the impact of encapsulation on the stability and bioactivity of peptides extracted from botanical sources: trends and opportunities.

Bioactive peptides derived from plant sources have gained significant attention for their potential use in preventing and treating chronic degenerative diseases. However, the efficacy of these peptides depends on their bioaccessibility, bioavailability, and stability. Encapsulation is a promising strategy for improving the therapeutic use of these compounds. It enhances their stability, prolongs their shelf life, protects them from degradation during digestion, and enables better release control by improving their bioaccessibility and bioavailability. This review aims to analyze the impact of various factors related to peptide encapsulation on their stability and release to enhance their biological activity. To achieve this, it is necessary to determine the composition and physicochemical properties of the capsule, which are influenced by the wall materials, encapsulation technique, and operating conditions. Furthermore, for peptide encapsulation, their charge, size, and hydrophobicity must be considered. Recent research has focused on the advancement of novel encapsulation methodologies that permit the formation of uniform capsules in terms of size and shape. In addition, it explores novel wall materials, including polysaccharides derived from unconventional sources, that allow the precise regulation of the rate at which peptides are released into the intestine.