Antimicrobial peptides (AMPs) are promising tools for combating microbial resistance. However, their therapeutic potential is hindered by two intrinsic drawbacks – low target affinity and poor in vivo stability. Macrocyclization, a process that improves the pharmacological properties and bioactivity of peptides, can address these limitations. As a result, macrocyclic peptides represent attractive drug candidates. Moreover, many drugs are macrocycles that originated from natural product scaffolds, suggesting that nature offers solutions to the challenges faced by AMPs. In this review, we explore natural cyclic peptides from the DBAASP database. DBAASP is a comprehensive repository of data on antimicrobial/cytotoxic activities and structures of peptides. We analyze the data on small (≤25 AA) ribosomal and non-ribosomal cyclic peptides from DBAASP according to their amino acid composition, bonds used for cyclization, targets they act on, and mechanisms of action. This analysis will enhance our understanding of the small cyclic peptides that nature has provided to defend living organisms.

For several millennia, doctors and healers used therapeutic ingredients extracted from minerals, plants, and animals. Among the active remedies derived from animals, the placenta was among the most popular ones. The use of placenta-based remedies has retained its significance in our time. The placenta-based therapeutics are indicated for stimulating immunity, wound healing, reduction of skin pigmentation, treatment of various patients suffering from gynecological disorders, etc. By the beginning of the 21st century, the majority of specific functions and chemical ingredients of the placenta were identified and described. However, only now active proteins and peptides with molecular weight of about ten kDa were revealed. Presumably, they play an essential role during the therapeutic application of placental extract. Hopefully, this discovery will start a new era in the research and clinical application of organotherapeutic remedies, including placenta.

Antimicrobial peptides (AMPs) are essential components of innate immunity across all species. AMPs have become the focus of attention in recent years as scientists are addressing antibiotic resistance, a public health crisis that has reached epidemic proportions. This family of peptides are a promising alternative to current antibiotics due to their broad-spectrum antimicrobial activity and tendency to avoid resistance development. A subfamily of AMPs interact with metal ions to potentiate their antimicrobial effectiveness, as such they have been termed metalloAMPs. In this work, we review the scientific literature of metalloAMPs that enhance their antimicrobial efficacy when combined with the essential metal ion, zinc (II). Beyond the role played by Zn(II) as a cofactor in different systems, it is well-known that this metal ion plays an important role in innate immunity. Here, we classify the different types of synergistic interactions between AMPs and Zn(II) into three distinct classes. By better understanding how each class of metalloAMPs uses Zn(II) to potentiate their activity, researchers can begin to exploit these interactions in the development of new antimicrobial agents and accelerate their use as therapeutics.

In 2020, the WHO established a road map for neglected tropical diseases 2021−2030 that aims to control and eradicate 20 diseases, including Leishmaniosis and Chagas disease. In addition, since 2015, the WHO has been developing a Global Action Plan on Antimicrobial Resistance. In this context, the achievement of innovative strategies as an alternative to replace conventional therapies is a first-order socio-sanitary priority, especially regarding endemic zoonoses in poor regions such as those caused by Trypanosoma cruzi and Leishmania spp infections. In this scenario, it is worth highlighting a group of natural peptide molecules (AMPs and CPPs) that are promising strategies for improving therapeutic efficacy against these neglected zoonoses, avoiding the development of toxicity and resistance of conventional treatments. This review presents the novelties of these peptide molecules and their ability to cross a whole system of cell membranes, as well as to stimulate the host immune defenses or even serve as a vector of molecules. The efforts of the biotechnological sector will make it possible to overcome the limitations of antimicrobial peptides through encapsulation and functionalization methods to obtain approval for these treatments to be used in clinical programs for the eradication of Leishmaniosis and Chagas disease.

Klebsiella pneumoniae is a multidrug-resistant opportunistic human pathogen related to various infections. As such, synthetic peptides have emerged as potential alternative molecules. Mo-CBP3-PepI has presented great activity against K. pneumoniae by presenting an MIC50 at a very low concentration (31.25 µg mL-1). Here, fluorescence microscopy and proteomic analysis revealed the alteration in cell membrane permeability, ROS overproduction, and protein profile of K. pneumoniae cells treated with Mo-CBP3-PepI. Mo-CBP3-PepI led to ROS overaccumulation and membrane pore formation in K. pneumoniae cells. Furthermore, the proteomic analysis highlighted changes in essential metabolic pathways. For example, after treatment of K. pneumoniae cells with Mo-CBP3-PepI, it was seen a reduction in the abundance of protein related to DNA and protein metabolism, cytoskeleton and cell wall organization, redox metabolism, regulation factors, ribosomal proteins, and resistance to antibiotics. These reductions lead to the inhibition of DNA repair, inhibition of cell wall turnover, protein turnover, and ROS accumulation leading to cell death. Our findings indicated that Mo-CBP3-PepI might have mechanisms of action against K. pneumoniae cells, mitigating the development of resistance and thus being a potent molecule to be employed in producing new drugs against K. pneumoniae infections.

PG16 is a broadly neutralizing antibody that binds to the gp120 subunit of the HIV-1 Env protein. The major interaction site is formed by the unusually long complementarity determining region (CDR) H3. The CDRH3 residue Tyr100H is known to represent a tyrosine sulfation site; however, this modification is not present in the experimental complex structure of PG16 with full-length HIV-1 Env. To investigate the role of sulfation for this complex, we modeled the sulfation of Tyr100H and compared the dynamics and energetics of the modified and unmodified complex by atomistic molecular dynamics simulations. Our results show that sulfation does not affect the overall conformation of CDRH3, but still enhances gp120 interactions both at the site of mutation and for the neighboring residues. This stabilization affects not only protein-protein contacts, but also the interactions between PG16 and the gp120 glycan shield. Further, we also investigated whether PG16-CDRH3 is a suitable template for the development of peptide mimetics. For a peptide spanning residues 93-105 of PG16 we obtained an experimental EC50 value of 3nM for the binding of gp120 to the peptide. This affinity can be enhanced by almost one order of magnitude by artificial disulfide bonding between residues 99 and 100F. In contrast, any truncation results in significantly lower affinity, suggesting that the entire peptide segment is involved in gp120 recognition. Their high affinity makes PG16-derived peptides useful building blocks for further optimization to obtain a potent inhibitor that efficiently blocks HIV-1 infection.

Urinary peptides are the products of systemic protein turn over which are excreted out of the body and hence can serve as an important biomarker for various pathophysiology. These peptides in other species of bovine have been reported to possess several bioactive properties. To investigate the potential of urinary peptides we generated a peptidome profile from the urine of Murrah Buffaloes (n=10). Urine samples were processed using <10 kDa MWCO filter, filtrate obtained was used for peptide extraction using Solid Phase Extraction (SPE). Eluate, obtained after SPE was extracted using ethyl acetate and the peptide-containing aqueous phase was collected. We extensively assessed the antimicrobial properties of the urinary aqueous phase. The nLC-MS/MS of the aqueous phase from 10 animals resulted in the identification of 8165 peptides originating from 6041 parent proteins. 76 Proteases responsible for the release of these sequences from precursor proteins were also identified. We further analyzed these peptide sequences to identify bioactive peptides and classify them into anti-cancerous, anti-hypertensive, anti-microbial, and anti-inflammatory groups with a special emphasis on antimicrobial properties. With this in mind, we simultaneously conducted experiments to evaluate antimicrobial properties of urinary aqueous extract on three pathogenic bacterial strains S. aureus, E. coli, & S. agalactiae.

The enormous influence in terms of bioactivity, affinity, and selectivity represented by the replacement of (L)-2,6-dimethyl tyrosine (Dmt) instead of Phenylalanine (Phe) into Nociceptin/orphanin (N/OFQ) neuropeptide analogues, has been well documented in literature. More recently, the non-natural amino acid (L)-2-methyl tyrosine (Mmt), with steric hindrance among Tyr and Dmt, has been studied because of the modulation of steric effects in opioid peptide chains. Here we report a new synthetic strategy to obtain Mmt based on the well-known Pd-catalyzed ortho-C(sp2)–H activation approach, since there is a paucity of other synthetic routes in literature, to achieve it. The aim of this work was to force only the mono ortho-methylation process over the double ortho-methylation one. In this regard we are pleased to report that the introduction of the dibenzylamine moiety on Tyr aromatic nucleus is a convenient and traceless solution to achieve such a goal. Interestingly, our method provided the aimed Mmt either as N-Boc or N-Fmoc derivatives ready to be inserted into peptide chains through solid phase peptide synthesis (SPPS). Importantly, the introduction of Mmt in place of Phe1 in the sequence of N/OFQ(1-13)-NH2 was very well tolerated in terms of pharmacological profile and bioactivity.

Blood-brain-barrier is a major obstacle in treating brain-related disorders as it does not allow to deliver drugs in the brain. In order to facilitate delivery of drugs in brain, we developed a method for predicting blood-brain-barrier penetrating peptides. These blood-brain barriers penetrating peptides (B3PPs) can act as therapeutic as well as drug delivery agents. We trained, tested, and evaluated our models on blood-brain-barrier peptides obtained from the B3Pdb database. First, we compute a wide range of peptide features then we select relevant peptide features. Finally, we developed numerous machine learning-based models for predicting blood-brain-barrier peptides using selected features. Our model based on random forest performed best on the top 80 selected features and achieved a maximum 85.08% accuracy with 0.93 AUROC. We also developed a web server, B3pred that implements our best models. It has three major modules that allow users to; i) predict B3PPs, ii) scanning B3PPs in a protein sequence, and iii) designing B3PPs using analogs. Our web server and standalone software is freely available at https://webs.iiitd.edu.in/raghava/b3pred/.

Peptides, both natural and synthetic, are well suited for a wide range of purposes and offer versa-tile applications in different fields such as biocatalysts, injectable hydrogels, tumor treatment, and drug delivery. The rising prominence of therapeutic peptides encompasses anticancer, antiviral, antimicrobial, and anti-neurodegenerative properties. The metals Na, K, Mg, Ca, Fe, Mn, Co, Cu, Zn, and Mo are ten of the twenty elements that are considered essential for life. Crucial for under-standing the biological role of metals is the exploration of metal-bound proteins and peptides. Aside from essential metals, there are other, non-essential metals that also interact biologically, exhibiting either therapeutic or toxic effects. Irregularities in metal binding contribute to diseases like Alzheimer’s, neurodegenerative disorders, Wilson's, and Menkes disease. Certain metal com-plexes have potential applications as radiopharmaceuticals. The examination of these complexes was done by preforming UV-Vis, IR, EPR, NMR spectroscopy, and X-ray analysis.

Coronaviruses are highly transmissible and pathogenic viruses for humans and animals. The vast quantity of information collected about SARS-CoV-2 during the pandemic helped to unveil details on the mechanisms behind the infection, which are still largely elusive. Recent research demonstrated that different class I/II human leukocyte antigen (HLA) alleles might define an individual susceptibility to SARS-CoV-2 spreading, contributing to the differences in the distribution of the infection through different populations; additional studies suggested that the homolog of the HLA in cats, the Feline Leukocyte Antigen (FLA), plays a pivotal role in the transmission of viruses. With these premises, this study aimed to exploit a bioinformatic approach for the prediction of the transmissibility potential of two distinct Feline Coronaviruses (FCoVs) in domestic cats (Feline enteric Coronavirus, FeCV and Feline Infectious Peritonitis Virus, FIPV) using SARS-CoV-2 as the reference model. We performed an epitope mapping of nonapeptides deriving from SARS-CoV-2, FeCV and FIPV glycoproteins and predicted their affinities for different alleles included in the three main loci in class I FLAs (E, H and K). The predicted complexes with the most promising affinities were then subjected to molecular docking and molecular dynamics simulations to provide insights into the stability and binding energies in the cleft. Results showed the FLA proteins encoded by alleles in FLA-I H (H*00501 and H*00401) and E (E*01001 and E*07001) loci are largely responsive to several epitopes deriving from replicase and spike proteins of the analyzed coronaviruses. The analysis of the most affine epitope sequences resulting from the prediction can stimulate the development of anti-FCoV immunomodulatory strategies based on peptide drugs.

In our modern days, macromolecular biomolecules are dethroning classical small molecule therapeutics because of improved targeting and delivery properties. Protamine – a small polycationic peptide represents such a promising candidate. In nature, it binds and protects DNA against degradation during spermatogenesis due to electrostatic interaction between the negatively charged DNA-Phosphate backbone and the positively charged protamine. Researchers are mimicking this technique in order to develop innovative nanopharmaceutical drug delivery systems, incorporating protamine as carrier for biologically active components such as DNA or RNA. The first key part of this review highlights ongoing investigation in the field of protamine-associated nanotechnology, discussing the self-assembling manufacturing process and nanoparticle engineering. Immune-modulating properties of protamine are referred which lead to the second key part protamine in novel vaccine technologies. Protamine-based RNA delivery systems in vaccines (some of them belong to the new class of mRNA-vaccines) against infectious disease and their use in cancer treatment are reviewed and an update on the current state of latest developments with protamine as pharmaceutical excipient for vaccines is given.

Alterations in myelin, the protective and insulating sheath surrounding axons, affect brain function, as is evident in demyelinating diseases where the loss of myelin leads to cognitive and motor dysfunction. Recent evidence suggests that changes in myelination, including both hyper- and hypo-myelination, may also play a role in numerous neurological and psychiatric diseases. Protecting myelin and promoting remyelination is thus crucial for a wide range of disorders. Oligodendrocytes (OLs) are the cells that generate myelin, and oligodendrogenesis (OLgenesis), the creation of new OLs, continues throughout life and is necessary for myelin plasticity and remyelination. Understanding the regulation of OLgenesis and myelin plasticity within disease contexts is therefore critical for the development of novel therapeutic targets. In our companion manuscript [1], we review literature demonstrating that multiple hormone classes are involved in the regulation of OLgenesis under physiological conditions. The majority of hormones enhance OLgenesis, increasing oligodendrocyte precursor cell differentiation and inducing maturation and myelin production in OLs. Thus, hormonal treatments present a promising route to promote remyelination. Here, we review literature on hormonal regulation of OLgenesis within the context of disorders. We focus on steroid hormones, including glucocorticoids and sex hormones, peptide hormones such as insulin-like growth factor 1, and thyroid hormones. For each hormone, we describe whether they aid in OL survival, differentiation, or remyelination, and we discuss their mechanisms of action, if known. Several of these hormones have yielded promising results in both animal models and in human conditions; however, a better understanding of hormonal effects, interactions, and their mechanisms will ultimately lead to more targeted therapeutics for myelin repair.

The brain’s capacity to respond to changing environments via hormonal signaling is critical to fine-tuned function. An emerging body of literature highlights a role for myelin plasticity as a prominent type of experience-dependent plasticity in the adult brain. Myelin plasticity is driven by oligodendrocytes (OLs) and their precursor cells (OPCs). OPC differentiation regulates the trajectory of myelin production throughout development, and importantly, OPCs maintain the ability to proliferate and generate new OLs throughout adulthood. The process of oligodendrogenesis (OLgenesis), the creation of new OLs, can be dramatically influenced during early development and in adulthood by internal and environmental conditions such as hormones. Here, we review the current literature describing hormonal regulation of OLgenesis within physiological conditions, focusing on several classes of hormones: steroid, peptide, and thyroid hormones. We discuss hormonal regulation at each stage of OLgenesis and describe mechanisms of action, where known. Overall, the majority of hormones enhance OLgenesis, increasing OPC differentiation and inducing maturation and myelin production in OLs. The mechanisms underlying these processes vary for each hormone but may ultimately converge upon common signaling pathways, mediated by specific receptors expressed across the OL lineage. However, not all of the mechanisms have been fully elucidated, and here, we note the remaining gaps in the literature, including the complex interactions between hormonal systems and with the immune system. In the companion manuscript in this issue [1], we discuss the implications of hormonal regulation of OLgenesis for neurological and psychiatric disorders characterized by white matter loss. Ultimately, a better understanding of the fundamental mechanisms of hormonal regulation of OLgenesis across the entire lifespan, especially in vivo, will progress both basic and translational research.

Small peptides (SPs), ranging from 5 to 100 amino acids, play integral roles in plants due to their diverse functions. Despite their low abundance and small molecular weight, SPs intricately regulate critical aspects of plant life, including cell division, growth, differentiation, flowering, fruiting, maturation, and stress responses. As vital mediators of intercellular signaling, SPs have garnered significant attention in plant biology research. This comprehensive review delves into SPs' structure, classification, and identification, providing a detailed understanding of their significance. Additionally, we summarize recent findings on the biological functions and signaling pathways of prominent SPs that regulate plant growth and development. The review also offers a forward-looking perspective on future research directions in peptide signaling pathways.

In the last 20 years, an increasing number of studies have been reported on membrane active peptides, which exert their biological activity by interacting with the cell membrane either to disrupt it and lead to cell lysis or to translocate through it to deliver cargos into the cell and reach their target. These peptides are attractive alternatives to currently used pharmaceuticals. Antimicrobial peptides (AMPs) and peptides designed for drug and gene delivery currently in the drug pipeline suggest that these membrane active peptides will soon constitute a significant percentage of the drug market. Here, we focus on two most prominent classes of membrane active peptides; AMPs and cell-penetrating peptides (CPPs). AMPs are a group of membrane active peptides that disrupt the membrane integrity or inhibit the cellular functions of bacteria, virus and fungi. CPPs are another group of membrane active peptides that mainly function as cargo-carriers even though they may also show antimicrobial activity to some extent. Biophysical techniques to understand how they interact with the membrane have shed light on the peptide–membrane interaction at various levels of detail. Structural investigation of membrane active peptides in the presence of the membrane provides important clues on the effect of the membrane environment on peptide conformations. Advances in live imaging techniques have allowed examination of peptide action at a single cell or single molecule level. In addition to these experimental biophysical techniques, molecular dynamics simulations provided clues on the peptide-lipid interactions and dynamics of the cell entry process at atomic detail. In this review, we summarize the recent advances in experimental and computational investigation of membrane active peptides with particular emphasis on two amphipathic membrane active peptides, the AMP melittin and the CPP pVEC.

Milk is newborn’s food and an emulsion full of all necessary components for neonatal growth. Its consumption is worldwide and is the base for all dairy products. Because of the latter, many new technologies are growing, among them proteomics; in order to give new insights in milk’s compounds and to maximize the beneficial potential for consumers’ health. In this review, we aimed to gather data of proteomics studies for the majority of dairy animals and elucidate the role of milk bioactive compounds. Furthermore, special reference was made to milk fat globule membrane (MFGM) peptides and the result of thermal treatment in milk proteins. Finally, the proteomic approach regarding adulterations was included in the review.

This systematic review and meta-analysis compile the in vitro antimicrobial efficacy of lactic acid bacteria (LAB) supernatant against three common dairy product pathogenic bacteria. After screening and analysis of full papers, identified by searches in PubMed, Scopus, and Web of Science databases, thirty-nine studies were regarded as relevant, and a total of 510 were extracted thereof. The effects of moderators, namely, pathogen’s concentration, susceptibility test method, agar type, incubation time and temperature, LAB supernatant pH and aliquot volume, on inhibition diameters were assessed by adjusting three pathogen-specific meta-regression models. Results showed that, in general terms, strains from the Enterococcus genus (15.90 ± 2.138 mm) displayed the highest inhibition values against L. monocytogenes, whereas Lacticaseibacillus strains were more effective against Staphylococcus aureus (11.89 ± 0.573 mm). Whereas the well diffusion test outperformed spot and disk diffusion tests; the pH of LAB supernatant resulted in high measurements of inhibition diameters (p

The antimicrobial peptide LRGG (LLRLLRRGGRRLLRLL-NH2) was designed and chemically synthesized in a study conducted by Jia et al. Gram-negative bacteria were found to be sensitive to LRGG and exhibited a high therapeutic index. Genetic engineering methods were used to create the prokaryotic fusion expression vector pQE-GFP-LRGG, and the resulting corresponding fusion protein GFP-LRGG was subsequently expressed and purified. The precursor GFP was then removed by TEV proteolysis, and pure LRGG was obtained after another round of purification and endotoxin removal. The prokaryotic-expressed antimicrobial peptide LRGG displays a broad-spectrum antibacterial effect on gram-negative bacteria and can eliminate 99.99% of E. coli within 100 minutes at the minimal MIC concentration. Compared to the chemically synthesized LRGG, the prokaryotic-expressed LRGG exhibits similar temperature, pH, salt ion, serum stability, and cell selectivity. Furthermore, prokaryotic-expressed LRGG showed excellent therapeutic effects in both the infection model of cell selectivity and no embryotoxicity in a Galleria mellonella infection model. The mechanism by which LRGG causes bacterial death was found to be disruption of Gram-negative cell membrane.

One of the main families of antimicrobial peptides (AMPs) derived from the skin secretions of Hylidae frogs are dermaseptins. Among them, dermaseptin S4 (DS4) is characterized by its broad spectrum of activity against bacteria, protozoa and fungi. This study determined the physicochemical properties of the native peptide DS4(1-28) and its derivatives [DS4(1-28)a and DS4(1-26)a] isolated from the skin of the frog Phyllomedusa sauvagii and examined their antimicrobial properties against two marine pathogenic bacteria (Vibrio harveyi and V. anguillarum). The results indicate that DS4(1-26)a has high bacteriicdal activity against the tested strains and low hemolytic activity (> 30 % lysis at the highest tested concentration of 100 µg mL-1) compared to the other two peptides. In addition, all three peptides affected the membrane and cell wall integrity of both pathogenic bacteria, causing leakage of cellular contents, with DS4(1-26)a having the most severe impact. These properties were corroborated by tansmission electron microscopy and by the variation of cations in their binding sites due to the effect of AMPs. These results suggest that DS4 and its derivatives, particularly the truncated and amidated peptide DS4(1-26)a, could potentially be effective in the treatment of infections caused by these marine pathogenic bacteria. Future experiments are required to validate the use of DS4 in vivo in the prevention of fish bacterial diseases.

The receptor tyrosine kinase (EphB4), which produces erythropoietin, was identified as a molecular target for cancer. We identified six peptides as potential inhibitors of EphB4 using an integrated approach combining online bioinformatics tools. According to PeptideRanker, CSM-peptides, SCMTHP, and AcPEP analysis, all peptides are effective against cancer, with the exception of LTAPGQATLPT and VEQNLVPGLK. Furthermore, Toxin-Pred predicts that none of the peptides are toxic, and that only VQLPGWRVFP and TMKGPPDTLQ are potentially allergenic using AllerTop server. PepSite2, HPEPDOCK, and MDockPeP servers were utilized to perform molecular docking, and PacDock was used to examine molecular interactions. The docking score (kcal/mol) range of six promising anticancer peptides against EphB4 was recorded as follows: VQLPGWRVFP (-240.72), FTKDDEWSCFPF (-201.74), SYLPPLSAEVTAK (-177.52), LTAPGQATLPT (-162.23), VEQNLVPGLK (-157.30), TMKGPPDTLQ (-149.67). The main bindings on the allosteric site to VQLPGWRVFP were with R706, S815, R819 and P843 on EphB4. Collectively, our results demonstrated that M. oleifera peptides ameliorate cancer by inhibiting EphB4, and they are promising candidates for the prevention and treatment of cancer.

Bioactive peptides are promising cosmetic active ingredients that can improve skin health and appearance. They exhibit a broad spectrum of activity, including anti–ageing, antioxidant, antimicrobial and anti–inflammatory. The aim of this study was to develop a safe, stable and efficacious environmentaly friendly ("green") emulsion using milk protein hydrolysate as a model active ingredient. Potential emulsions were formulated with biodegradable emollients, stabilised with a naturally-derived mixed emulsifier and prepared by cold process. They were evaluated for rheological behavior (continuous rotation and oscillation tests), physical stability (dynamic mechanical thermal analysis - DMTA test) and texture profiles, as well as for cytotoxic, antioxidant and antimicrobial effects. Rheological characterization revealed shear-thinning flow behaviour with yield point from continuous rotation tests and predominantly elastic character from oscillation (amplitude and frequency sweep) tests, with small structural change detected in the DMTA test. These results implied satisfactory rheological properties and good stability. Texture analysis revealed acceptable spreadability and substantivity of the emulsions. The protein hydrolysate showed antioxidant activity. The developed emulsions showed low antibacterial activity against selected microorganisms, but this was due to the action of preservatives, not peptides. All potential emulsions showed desirable safety profile. The results obtained provide the basis for the next stage of formulation development, i.e. in vivo efficacy tests.

Fungal infections pose a significant threat to human health, particularly in immunocompromised individuals. Due to several factors, such as the inappropriate use of antimicrobial drugs, the emergence of resistance in microorganisms, and increasing demand for less cytotoxic antifungal agents for immunocompromised patients, antifungal research has gained significant attention in recent years. Antifungal peptides, and among them, cyclic peptides, are a class of antifungal agents that have been under development since the 1960s. Using cyclic peptides as a promising approach for combating antifungal resistance in pathogenic fungi has gained significant attention in recent years. The identification of antifungal cyclic peptides from various sources has been possible due to the widespread interest in peptide research in recent decades. It is increasingly important to evaluate the narrow to broad-spectrum antifungal activity and the mode of action of synthetic and natural cyclic peptides for both synthesized and extracted peptides. This short review aims to highlight some of the antifungal cyclic peptides isolated from bacteria, fungi, and plants, taking into consideration the progress made in peptide research. Additionally, it provides a summary of synthetic cyclic peptides with antifungal activity and discusses the potential future of utilizing combinations of antifungal peptides from different sources. The review underscores the need for further exploration of the novel therapeutic applications of these abundant and diverse cyclic peptides.

The treatment of peptides has played an important role in clinical practice since the discovery of insulin therapy in the 1920s. Over 60 peptide drugs are approved in the United States (US and other regional mar-kets, and peptides continue to undergo drug discovery steadily. Peptide research and development has lev-eraged a wider range of structures known from other plant sources, via pharmacology and medicinal molecular biology, beyond its conventional focus on individual endogenous peptides. We build a comprehensive database of peptides that have met scientific studies with more than 150 constantly evolving peptides. Here we provide a simple overview of the peptide-based drug therapy environment, comprising evolutionary points of view, structural properties, operational thresholds, and explanation of the therapeutic area.

In this study, an extracellular metalloprotease from Pseudoalteromonas sp. H2 was purified and identified. The EH2 maintained more than 80% activity over a wide pH range of 5-10 and the stability was also nearly independent of pH. More than 65% of activity was detected in a wide temperature of 20-70℃. The high stability of the protease in presence of different surfactants and oxidizing agent was also observed. Moreover, we also investigated the antioxidant activities of the hydrolysates generated from porcine and salmon skins collagen by EH2. The results show that salmon skins collagen hydrolysates demonstrated higher DPPH (42.88%±1.85) and hydroxyl radical (61.83%±3.05) scavenging activity compared to those produced with porcine skins collagen. For oxygen radical absorbance capacity, the hydrolysates from porcine skins collagen had higher efficiency (7.72±0.13 μmol·TE/μmol). Even 1 nM mixed peptides can effectively reduce the levels of intracellular ROS. And the two type of substrates exerted the best antioxidant activity when hydrolyzed for 3 hours. Hydrolysis time and type of substrate exerted important effect on the antioxidant property of hydrolysates. This study may have implications for the potential application of marine protease in biocatalysis industry. In addition, the hydrolyzed peptides from meat waste containing beneficial collagen by protease have good antioxidant activity indicating it may be a potential additive in the food processing industry and cosmetics industry.

Background According to some authors, a single isolated measurement of serum BNP executed on hospital admission would not be a sufficiently accurate method to predict the outcome of patients with ADHF. Aims For verifying this assumption, a retrospective study was conducted on patients hospitalized for ADHF. Our main objective was to ascertain whether there was any difference in midterm mortality among patients with rising BNP at discharge as compared to those with decreasing BNP at discharge. Methods Medical records were examined so as to make a partition of the ADHF patient population into two groups, the former characterized by a rise in BNP during hospitalization, and the latter exhibiting a decrease in BNP in the measurement taken at hospital discharge. Results 177 patients were enrolled in a retrospective study. Among them, 53 patients (29.94%) had increased BNPs at the time of discharge, whereas 124 (70.06%) showed decreases in serum BNP during their hospital stay. The group with patients who exhibited BNP increases at the time of discharge had higher degree of congestion evident in the higher frequency of persistent jugular venous distention and persistent orthopnea at discharge. Moreover, patients with increased BNP at the time of discharge had a lower reduction in inferior vena cava maximum diameter [1.58 ± 2.2 mm vs. 6.32 ± 1.82 mm; p (one-way ANOVA)=0.001]. In contrast, there was no significant difference in weight loss when patients with increased BNP at discharge were compared to those with no such increase. A total of 14 patients (7.9%) died during the six-month follow-up period. Cox proportional hazard analysis revealed that BNP increase at the time of discharge was an independent predictor of six-month all-cause mortality after adjustment for age, sodium at discharge, creatinine at discharge and New York Heart Association (NYHA) class at discharge (hazard ratio 34.49; 95% confidence intervals: 4.55–261.06; P =0.001). Conclusions Among patients with history of ADHF, more elevated BNP levels at the time of discharge from the hospital compared to those detected at admission identify a patient subset with higher grade of congestion and higher six-month mortality.

Patients with hematologic malignancies or non-malignant diseases may undergo allogeneic hematopoietic cell transplantation (HCT), which represents a potential curative treatment. However, they are still at risk of life-threatening complications, such as relapse, acute graft-versus-host disease (GvHD), and opportunistic infections. These complications are more likely if T cell reconstitution is delayed during the initial 3-4 months after HCT. Therefore, it is of clinical importance to advance early peripheral T cell expansion.The thymus is the cradle of T cell production, but it is extremely sensitive to conditioning drugs used in HCT. As a result, egress of T cells from the thymus is abrogated during the first 3-6 months after HCT. Instead, early T cell reconstitution depends on peripheral expansion of engrafted donor T cells. However, besides its established function to produce T cells, the thymus also produces thymic peptides with hormone-like activity. These molecules play an essential part in the development and nature of immune responses, and may have a role in modulating T cell expansion and function after HCT. In this review, we investigate the role of thymic peptides in shaping the dynamics of immune reconstitution early after HCT. Furthermore, we summarize current reports on clinical application of thymic peptides post-HCT and discuss their potential use in improving patient outcomes.

Background/Objectives: Survival prospects following SARS-CoV-2 infection may extend beyond the acute phase, influenced by various factors including age, health conditions, and infection se-verity. However, this topic has not been studied in detail. We therefore investigated mortality risk post-acute COVID-19 in the CRIT-COV-U cohort. Methods: We could retrieve survival data from 651 patients and also assess the association between urinary peptides and future death. Data spanning until December 2023 were collected from six countries, comparing mortality trends with age- and sex-matched non-infected controls. A death prediction classifier was developed and validated using pre-existing urinary peptidomics datasets. Results: Notably, 13.98% of post-COVID-19 patients succumbed during the follow-up, with mortality rates significantly higher than non-infected controls, particularly evident in younger individuals (<65 years). These data for the first time demonstrate that SARS-CoV-2 infection highly significantly increases the risk of mortality not only during the acute phase of the disease, but also beyond, for a period of about one year. In our study we were further able to identify 201 Urinary peptides linked to mortality, and integrated these into a predictive classifier (DP201). Higher DP201 scores, alongside age and BMI, significantly predicted death. Conclusions: Our findings underscore the utility of urinary peptides in prognosticating post-acute COVID-19 mortality, offering insights for targeted interventions. Our data also suggest that mortality should be considered as one possible symptom or conse-quence of post-acute sequelae of SARS-CoV-2 infection, a fact that is currently overlooked.

Antimicrobial peptides (AMPs) have emerged as a promising class of bioactive molecules with the potential to combat infections associated with medical implants and biomaterials. This review article aims to provide a comprehensive analysis of the role of antimicrobial peptides in medical implants and biomaterials, along with their diverse clinical applications. The incorporation of AMPs into various medical implants and biomaterials has shown immense potential in mitigating biofilm formation and preventing implant-related infections. We review the latest advancements in biomedical sciences and discuss the AMPs that were immobilized successfully to enhance their efficacy and stability within the implant environment. We also highlight successful examples of AMP coatings for the treatment of surgical site infections (SSIs), Contact lenses, Dental Applications, AMP-Incorporated Bone Grafts, Urinary tract infections (UTIs), Medical Implants, etc. Additionally, we discuss the potential challenges and prospects of AMPs in medical implants, such as effectiveness, instability and implant-related complications. We also discuss strategies that can be employed to overcome the limitations of AMP-coated biomaterials for prolonged longevity in clinical settings.

Cruor, the main component responsible for the red color of blood in mammals, contains 90% hemoglobin, a protein that is considered a rich source of bioactive peptides. The aim of the present study is to evaluate the potential of human hemoglobin as a source of bioactive peptides, compared to bovine hemoglobin, which has been widely studied in recent years. More specifically, the study focused on neokyotorphin, a natural antimicrobial peptide and meat preservative, which corresponds to the α137-141 fragment of bovine hemoglobin produced by enzymatic hydrolysis. Firstly, the results of the in-silico analysis showed that the two types of hemoglobin have a high similarity for the α and β chains (identities of 88% and 91%, respectively). Secondly, both types of hemoglobin were subjected to similar enzymatic hydrolysis conditions, which showed that the hydrolysis of human hemoglobin followed the same reaction mechanism as the hydrolysis of bovine hemoglobin, the 'Zipper' mechanism. The hydrolysis conditions (23°C, pH 3.5) made it possible to obtain α137-141 in significant quantities to allow for significant valorization from an industrial point of view. The hydrolyses were able to take place at high initial substrate concentrations, allowing for faster and more efficient processing of the co-product in the same amount of time, with over 65% of this production achieved in just 30 min. Increasing the substrate concentration (from 1% to 10% (w/v)) resulted in a proportional increase in α137-141 production. Regardless of the concentrations tested, the kinetic pattern remained similar. Finally, the results of the mass spectrometry analysis revealed the presence of several bioactive peptides in cattle and humans. Although some were known before, new bioactive peptides were discovered in human hemoglobin, such as four antibacterial peptides (α37-46 PTTKTYFPHF; α36-45 FPTTKTYFPH; α137-141 TSKYR; α133-141 STVLTSKYR), four opioid peptides (α137-141 TSKYR; β31-40 LVVYPWTQRF; β31-37 LVVYPWT), an ACE inhibitor (β129-135 KVVAGVA), an anticancer agent (β33-39 VVYPWTQ) and an antioxidant (α137-141 TSKYR). These peptides have never been found in human hemoglobin before, according to our knowledge. These results highlight the potential of human hemoglobin as a source of bioactive peptides useful for the food or pharmaceutical industry.

Bacillus licheniformis produce several classes of antimicrobial substances which are mainly either peptides or proteins. Among of them bacteriocins - peptides or proteins of different structural composition including synthesized by bacteria ribosomally; non-ribosomally synthesized peptides and cyclic lipopeptides; exopolysaccharides. Different representatives of these classes act against Gram-positive, Gram-negative bacteria, fungal pathogens and amoebae cells. In this review, a detailed classification of antimicrobial substances produced by B. licheniformis based on their chemical structure and mode of the synthesis and activity is presented. For some (rather limited number) of secreted antimicrobials mechanism of their harmful effect on the target cells is established, however, for many of them it remains unknown. The antimicrobial activity for most substances was studied in vitro only, however some substances were characterized in vivo and are found practical applications in medicine and veterinary. The cyclic lipopeptides with surfactant properties are applied in industry. In this review, a special attention on antimycobacterials produced by B. licheniformis is made as a possible approach to combat multidrug resistant and latent tuberculosis. Indeed, a number of peptides and proteins revealed strong antimycobacterial activity. However, medical application of some bacteriocins with promising in vitro antimycobacterial activity is limited by their toxicity for animals and humans. In this connection, similarly with the enhancement of the antimycobacterial activity of natural bacteriocins using genetic engineering, reduction of the toxicity by the same approach looks feasible. A unique capability of B. licheniformis to synthesize and produce a bouquet of different antibacterial compounds allow to consider this organism as a universal natural vehicle for antibiotic substances in form of probiotic cultures strains to combat various types of pathogens including mycobacteria.

Cryptococcus neoformans is a human multidrug-resistant yeast with high mortality rates in immunocompromised patients. Recently, the synthetic peptide Mo-CBP3-PepII emerged as a potent anticryptococcal molecule with an MIC50 at low concentration. Here, the mechanisms of action of Mo-CBP3-PepII were deeply analyzed to provide new information about how it led C. neoformans cells to death. Light and fluorescence microscopies, analysis of enzymatic activities, and proteomic analysis were employed to understand the effect of Mo-CBP3-PepII on C. neoformans cells. Light and fluorescence microscopies revealed Mo-CBP3-PepII induced the accumulation of anion superoxide and hydrogen peroxide in C. neoformans cells. In addition to a reduction in the activity of superoxide dismutase (SOD), ascorbate peroxidase (APX), and catalase (CAT) in the cells treated with Mo-CBP3-PepII. In the presence of Ascorbic acid (AsA), no ROS were detected and Mo-CBP3-PepII lost the inhibitory activity against C. neoformans. Yet, Mo-CBP3-PepII inhibited the activity of lactate dehydrogenase (LDH), ergosterol biosynthesis, and induced the decoupling of cytochrome c from the mitochondrial membrane. Proteomic analysis revealed a reduction in the abundance of proteins related to energetic metabolism, DNA and RNA metabolism, pathogenicity, protein metabolism, cytoskeleton, and cell wall organization and division. Our findings indicated that Mo-CBP3-PepII might have multiple mechanisms of action against C. neoformans cells, mitigating the development of resistance and thus being a potent molecule to be employed in the production of new drugs against C. neoformans infections.

The plant immunity system is more and more revisited and new elements and roles are attributed to participate in the response to biotic stress. New terminology is also applied trying to identify different players in the whole scenario of immunity: Phytocytokines are one of those elements that are gaining more attention due the characteristics of processing and perception, and showing they are part of a big family of compounds that can amplify the immune response. This review aims to highlight the last findings about the role of phytocytokines in the whole immune response on biotic stress, including basal and adaptive immunity, and to expose the complexity in their action in plant perception and signaling events.

Nervous necrosis virus (NNV) has spread throughout the world, affecting more than 120 freshwater and marine fish species. While vaccination effectively prevents disease outbreaks, the difficulty of producing sufficient viruses using cell lines continues to be a significant disadvantage for producing inactivated vaccines. This study, therefore, explored the application of synthetic peptides as potential vaccine candidates for the prevention of NNV in Asian seabass (Lates calcarifer). Using the epitope prediction tool and molecular docking, three predicted immunogenic B cell epitopes (30-32 aa) derived from NNV coat protein were selected and synthesised, corresponding to amino acid positions 5 to 34 (P1), 133 to 162 (P2) and 181 to 212 (P3). All the predicted peptides interact with Asian sea bass’s MHC class II by docking. The antigenicity of these peptides was determined through ELISA and all peptides were able to react with NNV-specific antibodies. Subsequently, the immunogenicity of these synthetic peptides was investigated by immunisation of Asian seabass with individual peptides (30 μg/fish) and a peptide cocktail (P1+P2+P3, 10 μg each/fish) by intraperitoneal injection, followed by a booster dose at day 28 post-primary immunisation. There was a subset of immunised fish that were able to induce upregulation of CD4 in the head kidney and spleen. Importantly, antibodies derived from fish immunised with synthetic peptides reacted with whole NNV virions. Taken together, these findings indicate that synthetic linear peptides based on predicted B cell epitopes exhibited both antigenic and immunogenic properties, suggesting that they could be potential vaccine candidates for the prevention of NNV in fish.

Both ricin and R. communis agglutinin (RCA120), belonging to the type II ribosome-inactivating proteins (RIPs-Ⅱ), are derived from the seeds of castor bean plant. They share very similar amino acid sequences, but ricin is much more toxic than RCA120. It is urgently necessary to distinguish ricin and RCA120 in response to public safety. Currently, mass spectrometric assays are well established for unambiguous identification of ricin by accurate analysis of differentiated amino acid residues after trypsin digestion. However, diagnostic peptides are relatively limited for unambiguous identification of trace ricin, especially in complex matrices. Here, we demonstrate a digestion strategy of multiple proteinases to produce novel peptide markers for unambiguous identification of ricin. LC-HRMS was used to verified the resulting peptides, among which only the peptides with uniqueness and good MS response were selected as peptide markers. Seven novel peptide markers were obtained from tandem digestion of trypsin and endoproteinase Glu-C in PBS buffer. From the chymotrypsin digestion under reduction and non-reduction conditions, eight and seven novel peptides were selected respectively. Using pepsin under pH 1~2 and proteinase K digestion, 6 and 5 peptides were selected as novel peptide markers. In conclusion, the obtained novel peptides from the established digestion methods can be recommended for the unambiguous identification of ricin during the investigation of illegal use of the toxin.

Polydiacetylene (PDA) inserted in films or in vesicles have received increasing attention due to PDA property to undergo a blue-to-red colorimetric transition along with a change from non-fluorescent to fluorescent upon application of various stimuli. In this review paper, the principle of the detection of various microorganisms (bacteria: directly detected or detected through the emitted toxins or through their DNA, and viruses) and of antibacterial and antiviral peptides based on these responsive PDA vesicles are detailed. The obtained analytical performances, when vesicles are in suspension or immobilized, are given and compared to those of the responsive vesicles mainly based on the vesicle encapsulation method. Many future challenges are then discussed.

Peptide-based inhibitors represent a promising approach for the treatment of HIV-1, offering a range of potential advantages including specificity, low toxicity, and the ability to target various stages of the viral lifecycle. This review outlines the current state of research on peptide-based anti-HIV therapies, highlighting key advancements and identifying future research directions. Over the past few years, there has been significant progress in developing synthetic peptide-based drugs that target various stages of the viral life cycle, including entry and replication. These approaches aim to create effective anti-HIV therapies. Additionally, peptides have proven valuable in the development of anti-HIV vaccines. In the quest for effective HIV vaccines, discovering potent antigens and designing suitable vaccine strategies are crucial for overcoming challenges such as low immunogenicity, safety concerns, and increased viral load. Innovative strategies for vaccine development through peptide research are, therefore, a key focus area for achieving effective HIV prevention. This review aims to explore the strategies for designing peptides with anti-HIV activity and to highlight their role in advancing both therapeutic and preventive measures against HIV.

The rapid evolution of drug resistance is one of the greatest health issues of the 21st century. There is an alarming situation to find new therapeutic strategies or candidate drugs to tackle ongoing multi-drug resistance development. The marine environment is one of the prime natural ecosystems on Earth, the majority of which is still unexplored, especially when it comes to the microbes. A wide variety of bioactive compounds have been obtained from a varied range of marine organisms, however, marine bacteria-produced bacteriocins are still undermined. Owing to the distinct environmental stresses that marine bacterial communities encounter, their bioactive compounds frequently undergo distinct adaptations that confer on them a variety of shapes and functions, setting them apart from their terrestrial counterparts. The ribosomally-synthesized and posttranslationally modified peptides (RiPPs), bacteriocins are of special interest because of their variety in structure and diverse potential biological activities. Additionally, the gut microbiome of marine creatures are largely unexplored source of new bacteriocins with promising activities. There is a huge possibility of novel bacteriocins from marine bacterial communities that might come out as efficient candidates to fight against antibiotic resistance, especially in light of the growing pressure from antibiotic-resistant diseases and industrial desire for innovative treatments. The present review summarizes known bacteriocins from marine bacteria, evolutionary aspects, challenges, and huge possibilities of unexplored bacteriocins from marine bacterial communities.

Antimicrobial peptides (AMPs) are regarded as a promising alternative to traditional antibiotics in the face of ever-increasing resistance. Our research has focused on crotalicidin (Ctn), an AMP from a snake venom, and a fragment thereof, Ctn[15-34], with improved in vitro antimicrobial and anticancer activities and remarkable serum stability. As the retroenantio versions of both AMPs maintained favorable profiles, in this work we evaluate the in vivo efficacy of both the na-tive-sequence AMPs and their retroenantio counterparts in a murine infection model with Aci-netobacter baumannii. A significant reduction in bacterial levels is found in the mice treated with Ctn[15-34]. However, contrary to expectations, the retroenantio analogues either exhibit toxicity or lack efficacy when administered to mice.

TNF cytokine receptor TNFR1 protein plays an important role in immune response and autoimmune diseases. Previously, we demonstrated that the peptide of the innate immunity protein Tag7 (PGLYRP1), designated as 17.1, is able to perform the functions of a full-size Tag7 protein, namely, to inhibit the transmission of a signal through the TNFR1 receptor and activate this receptor in complex with the major heat shock protein Hsp70. In this study, we show that two peptides derived from 17.1 – 17.1A and 17.1B have different affinities to the TNFR1 receptor and the Hsp70 protein, and each of them is able to perform only one of the above functions: peptide 17.1A is only able to inhibit signal conduction through the TNFR1 receptor, and peptide 17.1B can only activate this receptor is in complex with Hsp70. Thus, it is possible to modulate the activity of TNFR1 receptor and further perform its specific inhibition or activation in the treatment of various autoimmune or oncological diseases.

Proteins, as gifts from nature, provide structure, sequence, and function templates for designing biomaterials. Here, we reported an engineered toolkit derived from a natural block copolymer, RfA1. RfA1 is composed of positively charged polyelectrolyte linker regions interspersed with highly conserved polyampholyte motifs. These linkers and motifs are constructional fragments and ready-to-use building blocks for synthetic design and construction. One functional and editable feature of RfA1 derivatives is their preferential distribution to cytoplasm or nucleoplasm, in a fragment-replication-determined manner. Based on this property, a prices spatiotemporal Tet-on demo was established, which effectively transports cargo peptides into nuclei at selective time points. Moreover, the functional homogeneities of either motifs or linkers were also verified, making them standardized building blocks for synthetic biology. In summary, this study provides a modularized, orthotropic and well-characterized toolkit for precise and spatiotemporal regulation of protein nucleocytoplasmic localization.

Pain is a natural bioassay for detecting and quantifying biological activities of venoms. The painfulness of stings delivered by ants, wasps, and bees can be easily measured in the field or lab using the stinging insect pain scale that rates the pain intensity from 1 to 4, with 1 being minor pain, and 4 being extreme, debilitating, excruciating pain. The painfulness of stings of 96 species of stinging insects and the lethalities of the venoms of 90 species was determined and utilized for pinpointing future promising directions for investigating venoms having pharmaceutically active principles that could benefit humanity. The findings suggest several under- or unexplored insect venoms worthy of future investigations, including: those that have exceedingly painful venoms, yet with extremely low lethality – tarantula hawk wasps (Pepsis) and velvet ants (Mutillidae); those that have extremely lethal venoms, yet induce very little pain – the ants, Daceton and Tetraponera; and those that have venomous stings and are both painful and lethal – the ants Pogonomyrmex, Paraponera, Myrmecia, Neoponera, and the social wasps Synoeca, Agelaia, and Brachygastra. Taken together, and separately, sting pain and venom lethality point to promising directions for mining of pharmaceutically active components derived from insect venoms.

Prostanoids modulate the pathogenesis of vascular diseases such as atherosclerosis, in which inflammation has an important role. It is well known that inducible Ciclooxygenase-2 (COX-2) is responsible for prostanoid production associated with inflammation. Angiotensin II may be implicated through the expression of COX-2 in the vascular wall. The purpose of this study was to examine in angiotensin II-stimulated adventitial fibroblasts the anti-inflammatory activity of different food peptides by inhibiting COX-2 expression, and the production of pro-inflammatory prostanoids. Fibroblasts from aorta of Sprague-Dawley rats were incubated with different food derived peptides followed by incubation with Angiotensin II. COX-2 expression was determined by western blot, transcriptional activity by luciferase assays and prostaglandin E2 by enzyme immunoassay. COX-2 expression was inhibited in the presence of Val-Pro-Pro (bovine β-casein 84–86), Arg-Asp-Ile-Leu-Asn-Gln (ovalbumin 84–89) and Tyr-Arg-Gly-Gly-Leu-Glu-Pro-Ile-Asn-Phe (ovalbumin 125–134). Angiotensin II-induced prostaglandin E2 production was also reduced by all the above-mentioned sequences. The incubation with ovalbumin-derived peptides displayed a significant reduction of COX-2 promoter activity compared to the stimuli with Angiotensin II in transiently transfected cells. These three sequences could potentially be used as functional food ingredients to reduce inflammation related to cardiovascular diseases.

Oral mucositis is most frequently a toxic effect of chemotherapeutic and/or radiotherapeutic treatment, resulting from complex multifaceted biological events involving DNA damage. The clinical manifestations have a negative impact on the life quality of cancer patients. Preventive measures and curative treatment of mucositis are still not well established. The glycine has anti-inflammatory, immunomodulatory and cytoprotective actions, being a potential therapeutic in mucositis. The objective was to evaluate the effects of glycine on the expression of collagen and growth factors, platelet and epidermal in oral mucositis. The mucositis of which was induced by the protocol of Sonis. 40 hamsters were used, divided into two groups: Group I- control; Group II- supplemented with 5% intraperitoneal glycine, 2,0 mg/g diluted in hepes. Histopathological sections were used to perform the immune-histochemical method, the evaluation collagen expression and the growth factors: EGF and PDGF. It was observed that the group supplemented with glycine higher amounts of collagen expression and predominance type of collagen I. The glycine group presented lower immunoexpression of the growth factors, EGF and PDGF. The group supplemented with glycine showed a marked healing process of the oral mucosite, demonstrated by the predominance of collagen type I and reduction of growth factors, EGF and PDGF.

One of the bottlenecks to bringing new therapies to the clinic has been a lack of vectors for delivering novel therapeutics in a targeted manner. Cell penetrating peptides (CPPs) have received a lot of attention and have been the subject of numerous developments since their identification nearly 3 decades ago. Known for their transduction abilities, they have generally been considered inert vectors. In this review we present a schema for their classification, highlight what is known about their mechanism of transduction, and outline the existing literature, as well as our own experience, vis a vis the intrinsic anti-inflammatory properties that certain CPPs exhibit. Given the inflammatory responses associated with viral vectors, CPPs represent a viable alternative to these vectors; further, the anti-inflammatory properties of CPPs, mostly through inhibition of the NF-κB pathway, are encouraging. Much work in relevant animal models, toxicity studies in large animal models, and ultimately human trials are needed before their potential is fully realized.

Background: Functional beverages are ordinary foods with components or ingredients able to provide specific health effects other than purely nutritional effect. Aim: This study aims to formulate a new functional beverage obtained from the recovery of YoAlp® whey, a bovine fermented milk obtained using autochthonous starter cultures, added with Renetta Canada/Golden Delicious-Raspberry (YWL), Renetta Canada-Aronia (YWA) and Ravèntse (YWR) fruit juices to improve the functionality of the beverage. Methods: Microbiological analysis and chemical characterization, by a proteomic approach and spectrophotometric microassay, have been conducted on beverages to investigate the expected shelf-life and potential health promoting effects. Results: The microbiological assessment confirmed the safety of the beverages and the probiotic bacteria vitality over a shelf life of seven days. Biochemical analysis performed highlight the presence of different bioactive peptides (ACE inhibitory and antioxidant), and the betacasomorphin 9 (BCM-9), marker of A2 variant of β-casein typically present in Aosta Valley cattle breeds. Moreover, high amounts of total polyphenols, which concur to the antioxidant activity values observed, have been found. Furthermore, ACE inhibitory activity is well correlated to the bioactive peptides detected. Finally, a consumer test confirmed the YoAlp® whey-based beverages appreciation. Conclusions: The production of these beverages could represent a huge opportunity for Aosta Valley’s farms to increase their income and competitiveness, creating a circular economy by recovering a by-product like whey and following sustainability and healthy trends, which are driving the food sector.

(1) Background: We comprehensively evaluated the expression of therapeutically targetable immune checkpoint molecules on celiac disease. We have focused on the alteration of the CD200/CD200R pathway and Elafin expression in celiac disease and discussed their roles in regulating the immune response. There are limited data related to the expression or function of these molecules in celiac disease. This finding could significantly contribute to the understanding of the clinical manifestation of celiac disease; (2) Methods: CD200, CD200R and Elafin distribution was determined by ELISA and immunohistochemistry analyses in serum and biopsies of CD patients. Analyses of Th1 and Th17 cytokines were determined. PCR amplification of a fragment of the PI3 gene was carried out using genomic DNA isolated from whole blood samples of the study subjects. Different aliquots of the PCR reaction product were subjected to RFLP analysis for SNP genotyping and detection; (3) Results: We characterized the expression and function of the CD200–CD200R axis and PI3 in celiac disease. A significantly higher level of soluble CD200, CD200R and lower expression of PI3 in serum of CD patients was observed compared to healthy controls. Consistent with our results, CD200 expression is regulated by IFN-gamma. Interaction of CD200/CD200R leads to production of type Th1 and Th17 cytokines. Regarding the PI3 genotype, the CT genotype proportion SNP rs1733103 and the GG genotype SNP rs41282752, were predominant in CD patients. SNP rs1733103 showed a significant association between the SNP variables and CD; (4) Conclusions: In celiac disease the immune checkpoint is compromised or dysregulated, which can contribute to inflammation and the autoimmunity process. The study of these checkpoint points will led to the development of targeted therapies aimed at restoring immunological balance in CD. Specific coding regions of the PI3 gene splice variants predispose the Elafin protein, both at the transcriptional and post-translational levels, to modify its expression and function, resulting in reduced differential functional protein levels in patients with active celiac disease.

Sporothrix brasiliensis is the most pathogenic species, responsible for the Brazilian cat-transmitted sporotrichosis hyperendemics. In this scenario, investigation of pathogen-host interaction can provide relevant information for future treatment strategies. To this end, the invertebrate Galleria mellonella has proven to be a suitable alternative to evaluate the virulence of pathogenic fungi since the insect immune system is similar to the mammalian innate immune response. The aim of this work was to investigate phenotypic and molecular aspects of the immune response of G. mellonella throughout the S. brasiliensis infection. Hemocyte density and the evolution of the fungal load were evaluated. In parallel, RT- qPCR expression analysis of genes encoding antimicrobial peptides (Gallerimycin and Galiomycin) and stress management genes (C7 Contig 15362 and C8 Contig 1910) was conducted. Fungal load and hemocyte densities were proportionally increasing simultaneously to the deleterious morphological events and larvae mortality. Gallerimycin, C7 Contig 15362, and C8 Contig 19101 genes were positively regulated (p<0.05) at distinct moments of S. brasiliensis infection, characterizing a time-dependent and alternately modulated profile. Galiomycin gene expression remained unchanged. Our results contribute to the future proposal of potential alternative pathways to treat and, consequently, control S. brasiliensis zoonosis, a major public Health in Latin America.

Spinal cord injury is a traumatic injury that causes a catastrophic state in patients due to neuronal deficits, loss of motor and sensory function, and this is due to secondary deleterious events subsequent to mechanical damage that expands the death of neural cells beyond the initial trauma. One of the most important events is inflammation, which activates molecules like pro-inflammatory cytokines (IL-1beta, IFN-gamma, TNF-alpha) that provoke a toxic environment, inhibiting axonal growth and exacerbating CNS damage. As there is no effective treatment, one of the strategies developed is neuroprotection to preserve healthy neural tissue and reduce neuroinflammation. Some of the neuroprotective treatments that have been practiced are the use of cell therapy, the administration of peptides and molecules or supplements that have been shown to favor an anti-inflammatory environment, and help to preserve the cells and tissues surrounding the injured area, favoring axonal growth and improving locomotor function. In this review we will explain some of these strategies used in different animal models of spinal cord injury, their activity as modulators of the immune system and the benefits they have shown.

Cryptococcus neoformans is a multidrug-resistant human pathogenic yeast responsible for infections in immunocompromised patients. Here, Itraconazole (ITR), a commercial antifungal drug with low effectiveness against C. neoformans, was combined with different synthetic peptides Mo-CBP3-PepII, RcAlb-PepII, RcAlb-PepIII, PepGAT, and PepKAA. The mechanisms of action responsible for the synergistic effect were evaluated for the best combinations by Fluorescence Microscopy (FM). The synthetic peptides enhanced the activity of ITR by 10-fold against C. neoformans. Our results demonstrated that the combinations could induce pore formation in the membrane and overaccumulation of ROS on C. neoformans cells. Our findings indicate that our peptides successfully potentialize the activity of ITR by reducing it by 10-fold to reach antifungal activity against C. neoformans. Therefore, synthetic peptides are potential molecules to act as co-adjuvant agents in treating Cryptococcal infections.

Cell-penetrating peptides (CPPs) have been evaluated as enhancers in drug delivery, their addition in medical formulations favors absorption allowing obtaining the pharmacological effect with lower drug doses. In vaccine formulations their inclusion has been also explored with interesting results. Currently mucosal vaccination constitutes a promising alternative with the main advantage of inducing both systemic and mucosal immune responses, which are crucial for control tumors and infections at mucosal tissues. The known CPP Penetratin was recently evaluated in vaccine formulations designed for nasal administration. The authors demonstrated that this non-covalent linked CPP could improve the antigen-specific systemic and mucosal antibody responses. In the present work we evaluate in Balb/C mice the nasal immune-enhancing effect of four CPPs. Animals were intranasally immunized with CPP and the recombinant hepatitis B surface protein (HBsAg) as model antigen. The IgG antibody response in sera and the mucosal IgA response were measured by ELISA. The IFN-g secretion response at spleen was also evaluated by ELISPOT and ELISA. Among the CPPs studied one novel peptide stand out by its ability to potentiate the humoral and cellular immune response against the co-administered antigen. Considering that the use of mucosal routes is a promising strategy in vaccination against infectious diseases and cancer, which are gaining special relevance nowadays in the development of novel candidates against SARS-CoV-2 and other potential emerging respiratory virus, the searching and development of safe mucosal adjuvants constitute a current need.

Background: In rheumatoid arthritis (RA), anti-citrullinated protein/peptide antibodies (ACPAs) are responsible for disease onset and progression, however, our knowledge is limited on ligand binding affinities of autoantibodies with different citrulline-peptide specificity. Methods: Citrulline-peptide specific ACPA IgGs were affinity purified and tested by ELISA. Binding affinities of ACPA IgGs and serum antibodies were compared by surface plasmon resonance (SPR) analysis. Bifunctional nanoparticles harboring a multi-epitope citrulline-peptide and a complement activating peptide were used to induce selective depletion of ACPA producing B cells. Results: KD values of affinity purified ACPA IgGs varies between 10-6-10-8 M and inversely correlate with disease activity. Based on their cross-reaction with citrulline-peptides we designed a novel multi-epitope peptide, containing Cit-Gly and Ala-Cit motifs in two-two copies, separated with a short, neutral spacer. This peptide detects antibodies in RA sera with 66 % sensitivity and 98 % specificity in ELISA and is recognized by 90% of RA sera, while none of the healthy samples in SPR. When coupled to nanoparticles, the multi-epitope peptide specifically targets and depletes ACPA producing B cells ex vivo. Conclusions: The unique multi-epitope peptide designed on the basis of ACPA cross-reactivity might be suitable to develop better diagnostics and novel therapies for RA.

Hydrogels evolved as an outstanding carrier material for local and controlled drug delivery that tend to the shortcomings of old conventional dosage forms for small drugs (NSAIDS) and large peptides and proteins. Aqueous swellable and crosslinked polymeric network structure of hydrogels is composed of various natural, synthetic and semisynthetic biodegradable polymers. Hydrogels have remarkable properties of functionality, reversibility, sterilizability, and biocompatibility. All these dynamic properties of hydrogels have increased the interest in their use as a carrier for peptides and proteins to be released slowly in a sustained manner. The therapeutical peptide and proteins are remarkable therapeutic agents in today’s world that allows the treatment of severe, chronic and life‐threatening diseases, such as diabetes, rheumatoid arthritis, hepatitis in an easy manner. Despite few limitations, hydrogels provide fine tuning of proteins and peptides delivery with enormous impact in clinical medicine. The primary objective of this article is to review current issues concerned with the therapeutics peptides and proteins and impact of remarkable properties of hydrogels on these therapeutic agents. Different routes for pharmaceutical peptides and proteins and superiority over other drugs candidates are presented. The article will also review literature concerning classification of hydrogels on different basis, polymers used, release mechanisms their physical and chemical characteristics and diverse applications.

Coffee processing generates a huge amount of waste which contains many natural products. Here, we report the discovery of a panel novel cell-penetrating and metal-ion binding microproteins designated coffeetide cC1a-c and cL1-6 from the husk of two popular coffee plants, Coffea canephora and Coffea liberica, respectively. Combining sequence determination and database search, we showed that the prototypic coffeetide cC1a is a 37-residue, eight-cysteine microprotein with a hevein-like cysteine motif but without a chitin-binding domain. NMR determination of cC1a revealed a compact structure that confers it resistance to heat and proteolytic degradation. Disulfide mapping together with chemical synthesis revealed that cC1a has a ginsentide-like, and not a hevein-like, disulfide connectivity. In addition, transcriptomic analysis showed that the 98-residue micrcoproten-like coffeetide precursor contains a three-domain arrangement like ginsentide precursors. Molecular modeling together with experimental validation revealed a Mg2+ and Fe3+ binding pocket at the N-terminus formed by three glutamic acids. Importantly, cC1a is amphipathic with a continuous stretch of 19 apolar amino acids, and which enables its cell penetration to target intracellular proteins, despite being highly negatively charged. Our findings suggest that coffee by-products could provide a source of ginsentide-like bioactive peptides that have potential to target intracellular proteins.

In recent years, bacterial resistance to conventional antibiotics has become a major concern in the medical field. The global misuse of antibiotics in clinics, personal use, and agriculture has accelerated this resistance, making infections increasingly difficult to treat and rendering new antibiotics ineffective more quickly. Finding new antibiotics is challenging due to the complexity of bacterial mechanisms, high costs and low financial incentives for the development of new molecular scaffolds, and stringent regulatory requirements. Additionally, innovation has slowed, with many new antibiotics being modifications of existing drugs rather than entirely new classes. Antimicrobial peptides (AMPs) are a valid alternative to small molecule antibiotics offering several advantages, including broad-spectrum activity and a lower likelihood of inducing resistance due to their multifaceted mechanisms of action. However, AMPs face challenges such as stability issues in physiological conditions, potential toxicity to human cells, high production costs, and difficulties in large-scale manufacturing. A reliable strategy to overcome the drawbacks associate with the use of small molecule antibiotics and AMPs is the combination therapy, namely the simultaneous co-administration of two or more antibiotics or the synthesis of covalently linked conjugates. This review is intended to cover and critically analyze the design, synthetic strategies, antibacterial activity and, when possible, the cytotoxicity of antibiotic-AMPs conjugates.

Milk serves as a crucial source of natural bioactive compounds essential for human nutrition and health. The gastrointestinal digestion of milk proteins releases various biologically active peptides that positively impact human health and enhance physiological performance. With the growing production of cheese, Greek yogurt, and other dairy products, the volume of unutilized acid whey is significantly increasing. Acid whey, containing almost all whey proteins and approximately one-quarter of casein, presents a valuable raw material for generating peptides with potential health benefits. These peptides exhibit properties such as antioxidant, antimicrobial, anti-inflammatory, anticarcinogenic, antihypertensive, antithrombotic, opioid, mineral-binding, and growth-stimulating activities, contributing to improved human immunity and the treatment of chronic diseases. Enzymatic hydrolysis of milk proteins has been developed as a method to produce these peptides, facilitating their use in food fortification. To incorporate whey into functional beverages, it undergoes fermentation by lactic acid bacteria with proteolytic activity. To specifically enrich these beverages with peptides exhibiting targeted bioactivities, it is essential to select lactic acid bacteria strains possessing the requisite protease profiles. Additionally, the sensory properties of the final product can be enhanced using starter cultures with diverse metabolic capabilities, even in the absence of added flavoring agents. Given the imperative to develop waste-free technologies and enhance the profitability of the dairy industry, there is a growing need to efficiently utilize and transform acid whey into a broad spectrum of valuable products and food additives. This approach is justified not only by the nutritional benefits of the bioactive components in whey but also by the commercial potential of safe and health-enhancing products.

Natural host defensins, also sometimes termed antimicrobial peptides, are evolutionarily conserved. They have been studied as antimicrobials, but some pharmaceutical properties, undesirable for clinical use, led to the development of synthetic molecules with constructed peptide arrangements and/or peptides not found in nature. The leading development currently is synthetic small molecule nonpeptide mimetics, whose physical properties capture the characteristics of the natural molecules, and share their biological attributes. We studied brilacidin, an arylamide of this type, for its activity in vitro against fungi (40 clinical isolates, 20 species) that the World Health Organization has highlighted as problem human pathogens. We find antifungal activity at low concentrations for many pathogens, which indicates further screening for activity, particularly in vivo, is justified to evaluate this compound, and other mimetics, as attractive leads for development of effective antifungal agents.

Antimicrobial peptides (AMPs) are essential components of the innate immune system, present across a spectrum of organisms. Their versatility lies in their ability to effectively inhibit a wide range of pathogens including bacteria, fungi, parasites, and viruses. As the challenge of antibiotic resistance escalates and concerns surrounding antibiotic use grow, AMPs have emerged as promising alternatives. Their applications span diverse domains such as medicine, agriculture, food production, animal husbandry, and aquaculture, primarily due to their efficacy against resistant pathogens, signifying their crucial role in combating infections. Beyond their antimicrobial properties, research illuminates their intricate involvement in the immune response network. These peptides impact crucial processes including cytokine release, chemotaxis, wound healing, angiogenesis and the activation of the adaptive immune system. This comprehensive review delves into the multifaceted realm of AMPs, encompassing their classification, varied applications across industries and notably, their significance in diverse disease conditions like infections, skin diseases, cancer, oral diseases, lung diseases and cardiovascular ailments. Special attention is directed towards current advancements in clinical applications of peptide-based therapeutics, offering invaluable insights into their potential future integration within clinical settings.

Membrane processes, such as microfiltration, ultrafiltration, and nanofiltration are increasingly used for various applications in both upstream and downstream processing. Membrane-based processes play a critical role in the field of separation/purification of biotechnological products, including protein production/purification. The possibility of using membranes to separate peptides from a chicken byproduct hydrolysate and the effect of the performed downstream processing on the DPP-IV inhibitory activity of MDCR (mechanical deboning chicken residue) has been investigated. The chicken byproduct hydrolysate was prepared by enzymatic hydrolysis followed by MF, UF, NF, and RO separation. The LC-OCD analysis confirmed that NF and RO would retain the bioactive peptides in the concentrate in comparison to MF and UF. Bioactivity was correlated with molecular weight distribution profiles and average molecular weights. Permeates after ultrafiltration showed an IC50 value of 0.75 mg/mL, comparable to other potent DPP-IV inhibitors derived from various food sources, and significantly more potent compared to the microfiltration sample, which showed an IC50 value of 1.04 mg/mL. The average molecular weight of the permeates calculated from the SEC chromatograms was 883 g/mol for UF and 1437 g/mol for MF. Of the four membranes studied, the UF membrane showed the best separation properties with respect to maximizing the yield and up-concentration of the bioactive peptides. Overall, UF was demonstrated to be a feasible technology for the removal of the undesired high molecular weight substances and up-concentration of small molecular weight bioactive peptides from chicken byproduct hydrolysate.

Traumatic brain injury (TBI) is among the leading causes of mortality and long-term impairment globally. TBI has a dynamic pathology encompassing a variety of metabolic and molecular events that occur in two phases, primary and secondary. An external forceful blow to the brain initiates the primary phase, which is followed by a secondary phase that involves the release of calcium ions (Ca2+) and the initiation of a cascade of inflammatory processes, including mitochondrial dysfunction, rise in oxidative stress, activation of glial cells, and damage to the blood-brain barrier (BBB), resulting in paracellular leakage. There is currently no FDA-approved drug for TBI, but existing approaches rely on delivering small and macromolecular treatments, which are severely constrained by the BBB, poor retention, off-target toxicity, and complex pathology of TBI. Therefore, there is a demand for innovative and alternative therapeutics with effective delivery tactics for diagnosis and treatment of TBI. Tissue engineering and use of biomaterials is one such alternative approach. With this approach, neuronal stem cell therapy is combined with synthetically generated tissue materials such as hydrogels, self-assembling peptides, and electrospun nanofibers, which may induce neurite outgrowth, differentiation of human neural stem cells, and nerve gap bridging in TBI. This review examines tissue engineering and the use of biomaterials as potential treatments for TBI, including their synthesis, mechanisms of action, and limitations. The review also discusses challenges facing tissue engineering and biomaterial technology including survival rate of transplanted stem cells and the development of biodegradable, biocompatible, and mechanically flexible biomaterials. A better understanding of the mechanisms and drawbacks of these novel therapeutic approaches will help guide the design of future TBI therapies.

This study focuses on the enzymatic hydrolysis of hemoglobin, the main component of cruor that gives blood its red color in mammals. The antibacterial and antioxidant potential of human hemoglobin hydrolysates were evaluated in comparison to bovine hemoglobin. The results showed strong antimicrobial activity of the peptide hydrolysates against six bacterial strains, independent of the initial substrate concentration level. The hydrolysates also showed strong antioxidant activity measured by four different tests. In addition, the antimicrobial and antioxidant activity of the human and bovine hemoglobin hydrolysates showed little or no significant difference, with only the concentration level being the determining factor in their activity. The results of the mass spectrometry study showed the presence of a number of bioactive peptides, the majority of which have characteristics similar to those mentioned in the literature. New bioactive peptides were also identified in human hemoglobin, such as antibacterial peptides PTTKTYFPHF (α37-46), FPTTKTYFPH (α36-45), TSKYR (α137-141) and STVLTSKYR (α133-141), as well as an antioxidant TSKYR (α137-141). According to these findings, human hemoglobin represents a promising source of bioactive peptides beneficial to the food or pharmaceutical industries.

New antimicrobial approaches are essential to counter antimicrobial resistance. The drug development pipeline is exhausted with emergence of resistance resulting in unsuccessful trials. The lack of an effective drug developed from the conventional drug portfolio has mandated the introspection into the list of potentially effective unconventional alternate antimicrobial molecules. Alternate therapies, that are clinically explicable forms include monoclonal antibodies, antimicrobial peptides, aptamers and phages. Clinical diagnostics optimizes the drug delivery. In the era of diagnostic-based applications, it is logical to draw diagnostic based treatment for infectious diseases. Selection criteria of alternate therapeutics in infectious disease include detection, monitoring of response and resistance mechanism identification. Integrating these diagnostic applications is disruptive to the traditional therapeutic development. The challenges and mitigation methods need to be noted. Applying the goals of clinical pharmacokinetics that include enhancing efficacy and decreasing toxicity of drug therapy this review analyses the strong correlation of alternate antimicrobial therapeutics in infectious diseases. The relationship between drug concentration and the resulting effect defined by the pharmacodynamic parameters are also analyzed. This review analyzes the perspectives of aligning diagnostic initiatives into the use of alternate therapeutics with a particular focus on companion diagnostic applications in infectious diseases.

Antimicrobial peptides (AMPs) are a promising alternative to antibiotics to combat drug resistance in pathogenic bacteria. However, the development of AMPs with high potency and specificity remains a challenge, and new tools to evaluate antimicrobial activity are needed to accelerate the discovery process. As a step toward direct prediction of the experimental minimum inhibitory concentration (MIC) of AMPs, we proposed MBC-Attention, a combination of a multi-branch CNN architecture and attention mechanism. Using a curated dataset of 3929 AMP against Escherichia coli, the optimal MBC-Attention model achieved an average Pearson correlation coefficient of 0.775 and an RMSE of 0.533 (log μM) in three independent tests of 393 sequences each. This results in a 5–12% improvement in PCC and 7–13% improvement in RMSE compared with RF and SVM models. Ablation studies confirmed that both attention mechanisms contributed to performance improvement.

Antibiotic resistance (ABR) is concerning issue due to its direct and indirect repercusions on public health, since decreased therapeutic effect of certain antibiotic to treatment complications that can cause death. There are several mechanism as to how ABR can be transferred from one microoorganisms to another, and many of them are dependant many environmental factors. The food supply chain is a environment in which ABR gene transfer can occur is multiple pathways, which generate concerns regarding food safety. Here, we summarize relevant mechanisms which are implied in ABR in food supply chain but also we are addressing routes of transmission and prevalence of ABR, implications on public health, and the application of new alternatives to antibiotics such as antimicrobial peptides, mainly bacteriocins, in order to countermeasure ABR.

Electrostatic interactions have a determining role in conformational and dynamic behavior of polyelectrolyte molecules [1]. In this study, anionic polyelectrolyte molecules, poly(glutamic acid) (PGA) and poly(aspartic acid) (PASA), in water solution with the most commonly used K+ or Na+ counterions were investigated using atomistic molecular dynamics (MD) simulations. Seven common force fields, AMBER99SB-ILDN, AMBER14SB, AMBER-FB15, CHARMM22*, CHARMM27, CHARMM36m and OPLS-AA/L, both with their native parameters and with the non-bonded fix (NBFIX) and electronic continuum corrections (ECC) to were studied. These corrections have bene introduced to correct for the problem of overbinding of ions to the charged groups of polyelectrolytes. Physical properties, such as molecular sizes, local structure and dynamics, were studied using two types of common counterions, potassium and sodium. The results show that in some cases, the macroion size and dynamics depend strongly on the models (parameters) for the counterions due to strong overbinding of ions and charged side chain groups. The local structures and dynamics are more sensitive on dihedral angle parameterization resulting in a preference for defined monomer conformations amd the type of correction used.

Staphylococcus aureus can develop resistance by mutation, tranfection or biofilm formation. Resistance was induced in S. aureus by growth in sub-inhibitory concentrations of ciprofloxacin for 30 days. The ability of the antimicrobials to disrupt biofilms was determined using crystal violet and live/dead staining. Effects on the cell membranes of biofilm cells was evaluated by measuring release of dyes and ATP and nucleic acids. S. aureus did not develop resistance to the AMPs but resistance increased to ciprofloxacin by 128 times after 30 passages. Only peptides reduced biofilms of ciprofloxacin resistant cells. The antibiofilm effect of melimine with ciprofloxacin was more (27%) than with melimine alone at 1X MIC (p < 0.001). Similarly, at 1X MIC the combination of Mel4 and ciprofloxacin produced more (48%) biofilm disruption than Mel4 alone (p < 0.001). Combinations of either of the peptides with ciprofloxacin at 2X MIC released  66 nM ATP, more than either peptide alone (p  0.005). At 2X MIC, only melimine in combination with ciprofloxacin released DNA/RNA which was 3 times more than released by melimine alone (p = 0.043). These results suggest the potential use of melimine and Mel4 with conventional antibiotics for the treatments of S. aureus biofilms.

Since their identification over twenty-five years ago, the plethora of cell penetrating peptides (CPP) and their applications has skyrocketed. These 5 to 30 amino acid long peptides have the unique property of breaching the cell membrane barrier while carrying cargoes larger then themselves into cells in an intact, functional form. CPPs can be conjugated to fluorophores, activatable probes, radioisotopes or contrast agents for imaging tissues, such as tumors. There is no singular mechanism for translocation of CPPs into a cell, and therefore, many CPPs are taken up by a multitude of cell types, creating the challenge of tumor specific translocation and hindering clinical effectiveness. Varying strategies have been developed to combat this issue and enhance their diagnostic potential by derivatizing CPPs for better targeting by constructing specific cell activated forms. These methods are currently being used to image integrin expressing tumors, breast cancer cells, human histiocytic lymphoma and protease secreting fibrosarcoma cells, to name a few. Additionally, identifying safe, effective therapeutics for malignant tumors has long been an active area of research. CPPs can circumvent many of the complications found in treating cancer with conventional therapeutics by targeted delivery of drugs into tumors, thereby decreasing off-target side effects, a feat not achievable by currently employed conventional chemotherapeutics. Myriad types of chemotherapeutics such as tyrosine kinase inhibitors, anti-tumor antibodies and nanoparticles can be functionally attached to these peptides leading to the possibility of delivering established and novel cancer therapeutics directly to tumor tissue. While much research is needed to overcome potential issues with these peptides, they offer a significant advancement over current mechanisms to treat cancer. In this review, we present a brief overview of the research leading to identification of CPPs with a comprehensive state of the art review on the role of these novel peptides in both cancer diagnostics as well as therapeutics.

The redox chemistry of copper(II) is strongly modulated by the coordination to amyloid-β peptides and by the stability of the resulting complexes. Amino terminal copper and nickel binding motifs (ATCUN) identified in truncated Aβ sequences starting with Phe4 show very high affinity for copper(II) ions. Herein, we study the oxidase activity of [Cu-Aβ4-x] and [Cu-Aβ1-x] complexes toward dopamine and other catechols. The results show that the CuII-ATCUN site is not redox-inert, the reduction of the metal is induced by coordination of catechol to the metal and occurs through an inner sphere reaction. The generation of a ternary [CuII-Aβ-catechol] species determines the efficiency of the oxidation, although the reaction rate is ruled by re-oxidation of the CuI complex. In addition to the N-terminal coordination site, the two vicinal histidines, His13 and His14, provide a second Cu-binding motif. Catechol oxidation studies together with structural insight from the mixed dinuclear complexes Ni/Cu-Aβ4-x reveal that the His-tandem is able to bind CuII ions independently of the ATCUN site, but the N-terminal metal complexation reduces the conformational mobility of the peptide chain, preventing the binding and oxidative reactivity toward catechol of CuII bound to the secondary site.

Heterocytous cyanobacteria are among the most prolific source of bioactive secondary metabolites, including anabaenopeptins (APTs). A terrestrial filamentous Brasilonema sp. CT11 collected in Costa Rica bamboo forest, as black mat was studied using a multidisciplinary approach: genome mining and HPLC-HRMS/MS coupled with bionformatic analyses. Herein, we report the nearly complete genome consisting 8.79 Mbp with a GC content of 42.4%. Moreover, we report on three novel tryptophane-containing APTs; anabaenopeptin 788 (1), anabaenopeptin 802 (2) and anabaenopeptin 816 (3). Further, the structure of two homologues, i.e., anabaenopeptin 802 (2a) and anabaenopeptin 802 (2b) was determined by spectroscopic analysis (NMR and MS). Both compounds were shown to exert weak to moderate antiproliferative activity against HeLa cell lines. This study also provides the unique and diverse potential of biosynthetic gene clusters and an assessment of the predicted chemical space yet to be discovered from this genus.

Xenorhabdus nematophila and Photorhabdus luminescens are entomopathogenic symbionts that produce several toxic proteins that can interfere with the immune system of insects. Here, we showed that outer membrane proteins (OMPs) could be involved as virulence factors during bacterial symbiont pathogenesis. Purified OMPs from bacterial culture were injected fifth instar larvae of Spodoptera exigua Hübner. Larvae were surveyed for fluctuations in total haemocyte counts (THC), granulocyte percentage (cellular defence), protease, phospholipase A2 (PLA₂), and phenoloxidase (PO) activities (humoral defence) at specific time intervals. Changes in the expression of the three antimicrobial peptides (AMPs), cecropin, attacin, and spodoptericin, were also measured. Larvae treated with both types of OMPs had more haemocytes than did the negative controls. OMPs of X. nematophila caused more haemocyte destruction than did the OMPs of P. luminescens. The OMPs of both bacterial species initially activated insect defensive enzymes post-injection, their activating fluctuated in different ways. Attacin, cecropin and spodoptericin were up-regulated by OMP injections more than in normal larvae. The expression of these three AMPs was maximal at four hpi with P. luminescens OMPs treatment. Expression of the three AMPs in X. nematophila treatment was irregular and lower than in the P. luminescens OMPs treatment. These findings provide insights into the role of OMPs of entomopathogenic nematode bacterial symbionts in countering the physiological defenses of insects.

Gluten is a complex mixture of storage proteins in cereals like wheat, barley and rye. Prolamins are the main components of gluten. Their high content in proline and glutamine makes them water-insoluble and difficult to digest in the gastrointestinal tract. Partial digestion generates peptide sequences which trigger immune responses in celiac and gluten-sensitive patients. Gluten detection in food is challenging because of the diversity, in various food matrices, of protein proportions and their immunogenicity. Attempts to develop standard reference materials have been unsuccessful. We present here a summary of recent studies reporting the detection of dominant Gluten Immunogenic Peptides (GIP) sharing epitopes presented in the α-gliadin 33-mer, the most important celiac disease-immunogenic sequence within gluten. GIP were not only detectable and quantifiable in very different kind of difficult to analyze food, but also in stool and urine of celiac patients on a supposedly gluten-free diet (GFD), providing the first simple and objective means to assess adherence to the GFD. Methods to specifically and sensitively detect the most active GIP in food and biological fluids are rational candidates may use similar analytical standard references for determination of the immunopathological risk of gluten exposure in gluten-related diseases.

Based on a new model of a possible origin of life, we establish an efficient and stable system undergoing structural reproduction, self-optimization and molecular evolution. This system is being formed under realistic conditions by the interaction of two cyclic processes, one of which offering vesicles as the structural environment, the other supplying peptides from a variety of amino acids as versatile building blocks. We demonstrate that structures growing in a combination of both cycles have the potential to support their own existence, to undergo chemical and structural evolution and to develop unpredicted functional properties. The key mechanism is the mutual stabilization of the peptides by the vesicles and of the vesicles by the peptides together with a constant production and selection of both. The development of the proposed system over time not only would represent one of the principles of life, but could also be a model for the formation of self-evolving structures ultimately leading to the first living cell. The experiment yields clear evidence on a vesicle-induced accumulation of membrane-interacting peptide which could be identified by liquid chromatography combined with high-resolution mass spectroscopy. We found that the selected peptide has an immediate effect on the vesicles, leading to i) reduced vesicle size, ii) increased vesicle membrane permeability, and iii) improved thermal vesicle stability.

Staphylococcus aureus bacteria are one of the leading microorganisms responsible for nosocomial infections as well as being the primary causative pathogen of skin and wound infections. Currently, the therapy of staphylococcal diseases faces many difficulties, due to a variety of mechanisms of resistance and virulence factors. Moreover, a number of infections caused by S. aureus is connected with biofilm formation that impairs effectiveness of the therapy. Short cationic lipopeptides that are designed on the basis of the structure of antimicrobial peptides structure are likely to provide a promising alternative to conventional antibiotics. Despite the fact that many research groups have proved a high antistaphylococcal potential of lipopeptides, the lack of unified protocols in determination of antimicrobial activity may be the reason of inconsistency of the results. The aim of this study was to learn how the use of various bacteriological media as well as solvents may affect activity of lipopeptides and their cyclic analogs.

The human androgen receptor (AR) is a ligand inducible transcription factor harboring an amino terminal domain (AR-NTD) hosting the ligand independent activation function. AR-NTD is intrinsically disordered and display aggregation properties conferred by the presence of a poly-glutamine (polyQ) sequence of 22 residues. The length of the polyQ sequence, as well as the presence of adjacent sequence motifs modulate this aggregation property. AR-NTD contains also a conserved sequence motif KELCKAVSVSM that displays an intrinsic property to form amyloid fibrils under mild oxidative conditions of its conserved cysteine residue. As peptide sequences with intrinsic ability to oligomerize are reported to have an impact on the aggregation of polyQ tract, we determined the effect of the KELCKAVSVSM on the polyQ stretch in the context of the AR NTD, using Atomic Force Microscopy (AFM). Here, we present evidence for a crosstalk between the amyloidogenic properties of the KELCKAVSVSM motif and the polyQ stretch at the AR NTD.

Amidst increasing awareness of diet-health relationships, plant-derived bioactive peptides are recognized for their dual nutritional and health benefits. This study investigates bioactive peptides released after Alcalase hydrolysis of protein from chachafruto (Erythrina edulis), a nutrient-rich South American leguminous plant, focusing on their behavior during simulated gastrointestinal digestion. Evaluating their ability to scavenge radicals, mitigate oxidative stress, and influence immune response biomarkers, the study underscores the importance of understanding peptide interactions in digestion. Methods included previous enzymatic hydrolysis with Alcalase to obtain peptides, and results show significant radical scavenging and oxidative stress protection by chachafruto peptides. These findings highlight the potential of chachafruto bioactive peptides in functional foods and nutraceuticals, supporting human health through dietary interventions.

The spike protein (S) of SARS-CoV-2 has been shown to bind to the human angiotensin-converting enzyme 2 (ACE2) receptor with much higher affinity compared to other coronaviruses. The binding interface between the ACE2 receptor plays a critical role in the entry mechanism of SARS-CoV-2 virus. There are specific amino acids involved in the interaction between S protein and ACE2 receptor. This specificity is critical for the virus to establish a systemic infection and cause COVID-19 disease. In the ACE2 receptor, the largest number of amino acids that play a crucial role in the mechanism of interaction and recognition with the S protein are located in the C-terminal part which represents the main binding region between ACE2 and S. This fragment is abundant in coordination residues such as aspartates, glutamates and histidine that could be targeted by metal ions. Zn²⁺ ion binds to the ACE2 receptor in its catalytic site and modulates its activity, but it could also contribute to the structural stability of the entire protein. The ability of the human ACE2 receptor to coordinate metal ions, such Zn²⁺, in the same region where it binds to the S protein could have a crucial impact in the mechanism of recognition and interaction of ACE2-S with consequences on their binding affinity that deserve to be investigated. To test this possibility, this study aims to characterize the coordination ability of Zn²⁺, and also Cu²⁺ for comparison, with selected peptide models of the ACE2 binding interface using spectroscopic and potentiometric techniques.

Paenibacilli are efficient producers of potent agents against bacterial and fungal pathogens, which are of great interest both for therapeutic applications in medicine as well as in agrobiotechnol-ogy. Lipopeptides produced by such organisms play a major role in their inactivation potential. In this work we investigated two lipopeptide complexes, the fusaricidins and the polymyxins, produced by Paenibacillus polymyxa strains DSM 32871 and M1 by MALDI-TOF mass spectrometry. The fusaricidins show potent antifungal activities and are distinguished by an unusual variabil-ity. For strain DSM 32871 we identified numerous yet unknown variants mass spectrometrically. DSM 32871 produces polymyxins of type E (colistins), while M1 forms polymyxins P. For both strains novel, but not yet completely characterized polymyxin species were detected, which pos-sibly are glycosylated. These compounds may be of interest therapeutically, because polymyxins attain increasing attention as last-resort antibiotics against multiresistant pathogenic Gram-negative bacteria. In addition, the volatilomes of DSM 32781 and M1 were investigated with a GC-MS approach using different cultivation media. Production of volatile organic com-pounds (VOCs) was strain and medium dependent. In particular, strain M1 manifested as an effi-cient VOC-producer that exhibited formation of 25 volatiles in total. A characteristic feature of Paenibacilli is the formation of volatile pyrazine derivatives.

Candida auris is considered an emerging Candida species that has rapidly spread all over the world. The evidence of its origin and emerging resistance is still unclear. The severe infection by this species causes significant mortality and morbidity among the elderly and immune compressed individuals. Development of drug resistance is the major factor associated with therapeutic failure of existing antifungal agents. Previous studies addressed the antifungal resistance profile and drug discovery for C. auris. However, comprehensive coverage of these information in a single investigation yet to be available. In this review, we have mainly focused on recent development in therapeutic strategy against C. auris. Based on the available information, several differential approaches were discussed, including existing antifungal drugs, chemical compounds, essential oils, natural products, antifungal peptides, immunotherapy, antimicrobial photodynamic therapy, drug repurposing, and drug delivery systems. Among them, chemical medications, natural products, and antifungal peptides are the prime contributors. However, a limited number of resources are available to prove the efficiency of these potential therapies in clinical usage. Hence, we hope that the data gathered in this review can encourage in vivo studies and clinical trials.

Background: We aimed to evaluate whether baseline GLS and NT-proBNP and their changes after cardiac resynchronization therapy (CRT) can predict the long-term clinical outcomes and the echocardiographic-based response to CRT (defined by 15% relative reduction in left ventricular end-systolic volume). Methods: We enrolled 143 patients with stable, ischemic heart failure (HF) undergoing CRT-D implantation. NT-proBNP and echocardiogram were obtained before and 6 months after. The patients were followed-up (median: 58 months) for HF-related deaths and/or HF hospitalizations (primary endpoint) or HF-related deaths (secondary endpoint) Results: 84 patients achieved the primary and 53 the secondary endpoint, while 104 patients were considered as CRT-responders and 39 as non-responders. At baseline, event-free patients had higher absolute GLS values (p&lt;0.001) and lower NT-proBNP serum levels (p&lt;0001) than those achieving primary endpoint. A similar pattern was observed in favor of CRT-responders vs non-responders. In Cox regression analysis, lower baseline absolute GLS value (HR=0.77;95%CI, 0.51-1.91; p=0.002), higher baseline NT-proBNP levels (HR=1.55;95%CI, 1.43-2.01;p=0.002) and diabetes (HR=1.27;95%CI, 1.12-1.98;p=0.003) were associated with lower incidence of primary endpoint. Conclusions: In HF patients undergoing CRT-D, baseline GLS and NT-proBNP concentrations may serve as prognostic factors, while they may predict the echocardiographic-based response to CRT.

The development of antimicrobial resistance to conventional antibiotics is a major global health challenge. Infections caused by multidrug-resistant gram-negative bacteria have been named one of the most urgent global health threats. Considerable efforts are devoted to developing new antibiotic drugs and investigating the mechanism of antibiotic resistance. Recently, Anti-Microbial Peptides (AMPs) have emerged as a new platform for the target and design of novel drug resistant anti-microbial agents promising a new therapeutic strategy. AMPs are rapid, potent, possess an unusually broad spectrum of activity, and have shown efficacy as topical agents. Unlike traditional therapeutics that interfere with essential bacterial enzymes, AMPs interact with microbial membranes through electrostatic interactions and physically damage cell integrity. However, naturally occurring AMPs have limited selectivity and modest efficacy. Therefore, recent efforts have focused on the development of synthetic AMP analogs as suitable drug targets. This work explores the development of novel antimicrobial agents which mimic the structure of graft-copolymers and mirror the mode of action of an AMP. Chitosan-graft-polypeptide side chains are synthesized by the ring-opening polymerization of N-carboxyanhydrides of L-lysine and L-leucine initiated from the functional groups of chitosan. The derivatives with random- and block-copolymer side chains are explored as drug targets. These graft copolymer systems exhibit activity against clinically significant pathogens and disrupt biofilm formation. This work highlights the potential of chitosan-graft-polypeptide structures in biomedical applications.

Artificially designed, chimeric peptide-based recombinant vaccines are novel approaches to combat the phylogenetically diverse Foot and Mouth Disease (FMD) Virus (FMDV) strains. Among seven distinct serotypes, only serotype O and A are dominantly circulating in Bangladesh and neighbouring countries of Asia, where transboundary transmission, recurrent outbreaks and emergence of novel lineages FMDV are highly prevalent. The objective of this study was to develop multi-epitope recombinant peptides, procuring immunogenicity against circulating diverse subtypes of FMDV serotype O and A. Two chimeric peptides, named B1 (41.0 kDa) and B3 (39.3 kDa), have been designed to incorporate potential B-cell and T-cell epitopes selected from multiple FMDV strains, including previously reported and newly emerged sub-lineages. After expression, characterization and immunization of guineapigs with considerable antigen load of B1 and B3 followed by the serological assays revealed the significant protective immunogenicity, developed from the higher (100 µg) doses of both antigens, against most of the currently prevalent serotype O and A strains of FMDV. The efficient expression, antigenic stability, and multivalent immunogenic potency of the chimeric peptides strongly indicate their credibility as novel vaccine candidates for FMDV serotypes O and A circulating in Bangladesh and surrounding territories.

(1) Background: The diphtheria toxoid antigen is a major component in pediatric and booster combination vaccines and is known to raise a protective humoral immune response upon vaccination. Although antibodies are considered critical for diphtheria protection, little is known about the antigenic determinants that maintain humoral immunity. (2) Methods: One hundred twelve 15-mer peptides covering the entire sequence of DTx protein were prepared by Spot-synthesis. Membrane-bound peptides immunoreactivity with sera from mice immunized with triple DTP vaccine allowed mapping of continuous B-cell epitopes, topological studies, MAPs synthesis, and ELISA development. (3) Results: Twenty epitopes were identified, being 2 in the signal peptide, 5 in the CD, 7 in the HBFT domain, and 5 in the RBD. Two 17-mer (CB/Tx-2/12 and CB/DTx-4-13) derived bi-epitope peptides linked by a Gly-Gly spacer were chemically synthesized. The peptides were used as antigens to coat ELISA plates and assayed with human (huVS) and mice vaccined sera (miVS) for in vitro diagnosis of diphtheria. The assay proved to be highly sensitive (99.96%) and specific (100%) either for huVS and miVS and when compared with a commercial ELISA test, demonstrate high performance. (4) Conclusions: Our work displayed the complete picture of the linear B cell IgG response epitope of the DTx responsible for the protective effect and demonstrated the specificity and eligibility to enter phase IIB studies of some epitopes to develop new and fast diagnostic assays.

Protein hydrolysates show great promise as bioactive food and feed ingredient and for valorization of side-streams from e.g. the fish processing industry. This study characterizes bulk emulsifying, foaming, and in vitro antioxidative properties of hydrolysates derived from cod frame by application of Alcalase and Neutrase, individually and sequentially as well as the influence of heat-treatment prior to hydrolysis. We present a novel approach that utilizes proteomics data for calculation of weighted mean peptide properties (length, molecular weight, and charge) and peptide-level abundance estimation. Using subsequent bioinformatic prediction of biofunctional properties to describe observed bulk properties, we are able to provide an in-depth hydrolysate characterization not previously seen. All hydrolysates displayed comparable or higher emulsifying activity and stability than sodium caseinate. Heat-treatment significantly increased stability but showed a negative effect on the activity and degree of hydrolysis. Combining peptide abundance with predicted emulsifying activity, we were able to identify several peptides that are likely linked to the observed differences in bulk emulsifying properties. In general, decreased hydrolysis resulted in significantly higher chelating activity, while the opposite was observed for radical scavenging activity. The study highlights the prospects of applying proteomics and bioinformatics for hydrolysate characterization and in food protein science.

Due to its similarity with human milk and its low allergenic properties, donkey milk has long been used as an alternative for infants and patients with cow's milk protein allergy. In addition, this milk is attracting growing interest in human nutrition because of presumed health benefits. It has antioxidant, antimicrobial, antitumoral, antiproliferative, and antidiabetic activity. Also, it stimulates the immune system, regulates the gastrointestinal flora, and prevents inflammatory diseases. Although all components of donkey milk can contribute to functional and nutritional effects, it is generally accepted that the whey protein fraction plays a major role. The purpose of this review is to highlight the active proteins and peptides of donkey milk in comparison with other types of milk, emphasizing their properties and their roles in different fields.

One of the most commonly known chronic neurodegenerative disorders, Alzheimer’s disease (AD), manifests the common type of dementia in 60–80% of cases. From a clinical standpoint, a patent cognitive decline and a severe change in personality, as caused by a loss of neurons, is~usually evident in AD with about 50 million people affected in 2016. The disease progression in patients is distinguished by a gradual plummet in cognitive functions, eliciting symptoms such as memory loss, and eventually requiring full-time medical care. From a histopathological standpoint, the~defining characteristics are intracellular aggregations of hyper-phosphorylated tau protein, known as neurofibrillary tangles (NFT), and depositions of amyloid β-peptides (Aβ) in the brain. The~abnormal phosphorylation of tau protein is attributed to a wide gamut of neurological disorders known as tauopathies. In addition to the hyperphosphorylated tau lesions, neuroinflammatory processes could occur in a sustained manner through astro-glial activation, resulting in the disease progression. Recent findings have suggested a strong interplay between the mechanism of Tau phosphorylation, disruption of microtubules, and synaptic loss and pathology of AD. The mechanisms underlying these interactions along with their respective consequences in Tau pathology are still ill-defined. Thus, in this review: (1) we highlight the interplays existing between Tau pathology and AD; and (2) take a closer look into its role while identifying some promising therapeutic advances including state of the art imaging~techniques.

(RADA)4 nanoscaffolds are excellent candidates for use as peptide delivery vehicles: they are relatively easy to synthesize with custom bio-functionality, and assemble in situ to allow a focal point of release. This enables (RADA)4 to be utilized in multiple release strategies by embedding a variety of bioactive molecules in an all-in-one ‘construct’. One novel strategy focuses on the local, on-demand release of peptides triggered via proteolysis of tethered peptide sequences. However, the spatial-temporal morphology of self-assembling nanoscaffolds may greatly influence the ability for enzymes to both diffuse into as well as actively cleave substrates. Fine structure and its impact on overall affect on peptide release is poorly understood. In addition, fractal networks observed in nanoscaffolds are linked to the fractal nature of diffusion in these systems. Therefore, matrix morphology and fractal dimension of virgin (RADA)4 and mixtures of (RADA)4 and matrix metalloproteinase 2 (MMP-2) cleavable substrate modified (RADA)4 were characterized over time. Sites of high (GPQG+IASQ, CP1) and low (GPQG+PAGQ, CP2) cleavage activity were chosen. Fine structure was visualized using established according to established methods. After 2 hrs of incubation, nanofiber networks showed an established fractal nature, however nanofibers continued to bundle in all cases as incubation times increased. It was observed that despite extensive nanofiber bundling after 24 hrs of incubation time, the CP1 and CP2 nanoscaffolds were susceptible to MMP-2 cleavage. The properties of these engineered nanoscaffolds characterized herein illustrate that they are an excellent candidate as an enzymatically initiated peptide delivery platform.

Current research in the field of antimicrobials is focused on the development of novel antibiotics and antimicrobial agents to counteract the huge dilemma that the human population is mainly facing in regards to the rise of bacterial resistance and biofilm infections. Host Defense peptides (HDPs) are a promising group of molecules for antimicrobial development as they share unique characteristics suitable for antimicrobial activity including their broad spectrum of activity and potency against bacteria. AamAP1 is a novel HDP that was identified through molecular cloning from the venom of the North African scorpion Androctonus amoeruxi. In vitro antimicrobial assays revealed that the peptide displays moderate activity against different strains of Gram-positive and Gram-negative bacteria. Additionally, the peptide proved to be highly hemolytic and displaying significantly high toxicity against mammalian cells. In our study, a novel synthetic peptide analogue named A3 was designed from the naturally occurring scorpion venom host defense peptide. The design strategy depended on modifying the amino acid sequence of the parent peptide in order to increase its net positive charge, percentage helicity and optimize other physico-chemical parameters involved theoretically in HDPs activity. Accordingly, A3 was evaluated for its in vitro antimicrobial and anti-biofilm activity individually and in combination with four different types of conventional antibiotics against clinical isolates of multi-drug resistant (MDR) Gram-positive bacteria. A3 was also evaluated for its cytotoxicity against mammalian cells. A3 displayed potent and selective in vitro antimicrobial activities against a wide range of MDR Gram-positive bacteria. Our results also showed that combining A3 with conventional antibiotics displayed a synergistic mode of action which resulted in decreasing the MIC value for A3 peptide as low as 0.125 µM. These effective concentrations were associated with negligible toxicities on mammalian cells. In conclusion, A3 exhibits enhanced activity and selectivity when compared with the parent natural scorpion venom peptide. The combination of A3 with conventional antibiotics may be pursued as a potential novel treatment strategy against MDR and biofilm forming bacteria.

It was previously reported that the peptide derived from LfcinB, R-1-R exhibited anti-Candida activity and this activity was enhanced in combination with an extract from the Bidens pilosa plant. However, the mechanism of action has not been studied. In the current study, a proteomic study was carried out, followed by a bioinformatic analysis and biological assays in both SC5314 strain and a fluconazole-resistant isolate of Candida albicans, after incubation with R-1-R. The proteomic data showed that after treatment with R-1-R in the two study strains, most of the differentially expressed proteins in comparison to controls were up-regulated. These proteins were mainly involved in membrane and cell wall biosynthesis, membrane transporters, oxidative stress, the mitochondrial respiratory chain, and response to DNA damage. Besides, the proteomic analysis of C. albicans parental strain SC5314 treated with R-1-R in combination with an ethanolic extract of B. pilosa was also undertaken. The proteins differentially expressed after this treatment were involved in similar functional processes as the proteins differentially expressed after treatment with R-1-R peptide alone, but at different levels and were mostly down-regulated. The biological assays allowed validation of the proteomic results, evidencing damage to the cell surface, generation of reactive oxygen species and decrease in mitochondrial membrane potential. The results provide information to begin to unlock the understanding of the complex antifungal mode of action of the R-1-R peptide, its combination with the extract, and possibly other derivatives of natural products.

Background/Aim: Stool gluten immunogenic peptides (GIP) determination is now considered a valuable tool for detecting dietary lapses in treated celiac disease (CeD) patients. Our study aims to explore the diagnostic performance of a qualitative lateral flow immunoassay point-of-care test (LFIA/PoCT) compared with a qualitative enzyme-linked immunosorbent assay (ELISA) for detection of stool GIP. Methods: The study is a post hoc analysis of stool samples collected in the real-life conditions from a cohort of gluten-free diet-treated CeD patients. Stool GIP was measured in µg/g of stool using an ELISA kit (iVYLISA GIP S®, Biomedal SL). For the qualitative evaluation of stool GIP, a LFIA/PoCT test (GlutenDetect®; Biomedal S.L) was employed, which reported results as positive or negative. The LFIA/PoCT performance was evaluated against ELISA, considered as the gold standard. GIP concentrations

Background: Fermented milks are a source of bioactive peptides with different potential benefits on human health (mainly antioxidant and antihypertensive effects) and may be considered as functional foods. Furthermore, those made with sheep milk have higher nutritional value and higher concentrations of proteins, fats, minerals, and vitamins, if compared to milk of other species. Objectives: Aim of this study was to perform a biochemical characterization and a comparison between sheep fermented milk (SFM) made with commercial starter cultures and sheep milk YoAlp®, a fermented milk obtained using local strains of lactic acid bacteria (LABs) (Streptococcus thermophilus and Lactobacillus delbrueckii) which have been isolated during years in Aosta Valley mountain dairy cattle farm. Methods: Sheep milk and fermented milk (Sarda and Lacaune breeds) have been collected and immediately analyzed. Biochemical characterization, by a proteomic approach, GC/MS and microtiter plate assay methods, have been conducted to evaluate their peptide, fatty acid and aromatic profile, and to assess potential health promoting effects. Results: The comparison between SFM and YoAlp® peptide profile shows a higher number of peptides using autochthonous starter cultures (52) than commercial starter cultures (35). Among these peptides 20,78% and 29,87%, respectively, are supposed to be potentially bioactive (DPP - dipeptidyl peptidase-IV - inhibitor, ACE - angiotensin converting enzyme – inhibitor and antioxidant). In both products, the fatty acids profile was similar to that of origin sheep milk; concerning aromatic profile, YoAlp® shows yogurt typical aromatic assets, similar to the commercial one. As far as bioactivity is concerned, ACE inhibitor activity is high for both samples. Similar values, as expected by peptide profile analysis, have been obtained using commercial and autochthonous starters. Even in the case of antioxidant capacity, peptide profile bioactivity prediction has been confirmed by the assay showing a DPPH inhibition higher for SFM than for YoAlp®, but this difference is not statistically significative. Conclusions: Local strains of lactic acid bacteria from Aosta Valley seems to work as well as the commercial, preserving biodiversity and typicality. However, further analyses are needed to identify peptides generated by autochthonous LABs in order to understand microbial proteolytic activities and to investigate gastric digestion resistance of bioactive peptides which can have positive effects on human health.

Despite great strides in diagnosis and treatment, cardiac diseases remain the number one cause of mortality in the developed world, with rates catching up at an alarming rate in the developing world. Targeted delivery of therapeutics, specifically to cardiomyocytes, would open up new frontiers. Our prior work using a combinatorial in vitro and in vivo phage display methodology identified a 12-amino acid long peptide that targets cardiomyocytes selectively after an intravenous injection in as little as 5 mins, and hence was termed cardiac targeting peptide (CTP: APHLSSQYSRT). CTP has been used to deliver imaging agents, small drug molecules, photosensitizing nanoparticles, exosomes, and even miRNA to cardiomyocytes. As a natural extension for development of CTP as a clinically viable cardiac vector, we now present toxicity studies performed on the peptide. In vitro viability studies were performed in a human left ventricular myocyte cell line with 10µM of Cyanine-5.5 labeled CTP (CTP-Cy5.5). In vitro ion channel profiles were completed for CTP followed by more extensive studies performed by Eurofins in stably transfected cell lines for an extensive list of GPCR-coupled receptors. Positive data for GPCR-coupled receptors was interrogated further with RT-qPCRs performed on mouse heart tissue. In vivo studies consisted of pre- and post-blood pressure monitoring acutely after a single CTP (10 mg/Kg) injection. Further in vivo toxicity studies consisted of injecting 60, 6-week-old, wild-type CD1, male/female mice (1:1) injected with CTP (150 µg/Kg) intravenously. Mice were weighed immediately before injections and daily thereafter. Cohorts of mice were euthanized on day 0, 1, 2, 7 and 14 with inhalational CO2, followed by opening of the chest cavity, collection of blood via cardiac puncture, complete blood count analysis, metabolic profiling, and finally, liver, renal and thyroid studies. Lastly, mouse cardiac MRI was performed immediately before and after injections of CTP (150 µg/Kg) to assess changes in cardiac size or function. Human left ventricular cardiomyocytes showed no decrease in viability after a 30 min incubation with CTP-Cy5.5. In the Eurofins Cardiac Profiler Qube, no significant activation or inhibition of any of seventy-eight protein channels was observed other than OPRM1 and COX2 at the highest tested concentration, neither of which were expressed in mouse heart tissue as assessed by RT-qPCR. There was no change in blood pressures before and after injections with CTP at 10 mg/Kg. Blood counts and chemistries showed no evidence of significant hematological, hepatic or renal toxicities. Lastly, there was no significant difference in cardiac function, size or mass before and after CTP injections. Our studies with CTP showed no activation or inhibition of GPCR-associated receptors in vitro. We found no signals indicative of toxicity in vivo. Most importantly, cardiac functions remained unchanged acutely in response to CTP uptake. Further studies, in good laboratory practices fashion, are needed with more prolonged, chronic administration of CTP conjugated to a specific cargo of choice before human studies can be contemplated.

Despite significant strides in prevention, diagnosis, and treatment, cardiovascular diseases remain the number one cause of mortality in the United States, with rates climbing at an alarming rate in the developing world. Targeted delivery of therapeutics to the heart has been a lofty goal to achieve with strategies ranging from direct intra-cardiac or intra-pericardial delivery, intra-coronary infusion, to adeno-associated viral vectors which have preference, if not complete cardio-selectivity, for cardiac tissue. Cell penetrating peptides (CPP) are 5-30 amino acid long peptides that are able to breach cell membrane barriers while carrying cargoes up to several times their size, in an intact functional form. Identified nearly three decades ago, the first of these CPPs came from the HIV coat protein transactivator of transcription. Although a highly efficient CPP, its clinical utility is limited by its robust ability to cross any cell membrane barrier, including crossing the blood-brain barrier and transducing neuronal tissue non-specifically. Several strategies have been utilized to identify cell or tissue specific CPPs, one of which is phage display. Using this latter technique, we identified a cardiomyocyte targeting peptide (CTP) more than a decade ago, a finding that has been corroborated by several independent labs across the world that have utilized CTP for myriad different purposes in pre-clinical animal models. The goal of this publication is to provide a comprehensive review of the identification, validation, and application of CTP, and outlining its potential in diagnostic and therapeutic applications especially in the field of targeted RNA interference.

Enzymatic hydrolysis of bovine and human hemoglobin yields a diversity of bioactive peptides, some of which are recognized for their antimicrobial properties. Certain small antimicrobial and anticancer peptides target protein synthesis inhibition. This study aims to produce bioactive peptides through enzymatic hemoglobin hydrolysis and evaluate their effects on cell growth inhibition, potentially serving as candidates for biological anticancer drugs. Rapid screening of candidate anticancer peptides derived from enzymatic hydrolysis of bovine and human hemoglobin was conducted using the Lepidium sativum radicle growth technique. The results demonstrate that: 1) the degree of hydrolysis (DH) significantly influences the production of bioactive peptides. DH levels of 3-10% yield considerably stronger radicle growth inhibition compared to DH 0 (native hemoglobin form), exhibiting three to four times greater intensity. 2) Certain peptide fractions from bovine hemoglobin exhibit higher activity than those from human hemoglobin. 3) The structural characteristics of peptides play a crucial role in the observed biological effects. 4) The α137-141 peptide is the most active among the fractions obtained from bovine (IC₅₀ = 29 ± 1 µg/mL) and human hemoglobin (IC₅₀ = 48 ± 2 µg/mL), standing out 10 to 15 times more potent than other hemoglobin fractions. In conclusion, these findings affirm the bioactivity of peptide fractions obtained from bovine and human hemoglobin. The study highlights how enzymatic hydrolysis of bovine and human hemoglobin can yield peptides that inhibit protein synthesis. Ultimately, these results suggest the feasibility of obtaining candidate peptides for biological anticancer drugs from both human and bovine hemoglobin sources. These scientific advances show new hope in the fight against cancer which affects a large number of people around the world.

Using density functional theory (DFT) and the information-theoretic approach (ITA) quantities to appreciate the energetics and properties of biopolymers is still an unaccomplished and ongoing task. To this end, we studied the building blocks of nucleic acid base pairs and small peptides. For base pairs, we have dissected the relative importance of energetic components by using two energy partition schemes in DFT. Our results convincingly show that the exchange-correlation effect predominantly governs the molecular stability of base pairs while the electrostatic potential plays a minor but indispensable role, and the steric effect is trivial. Furthermore, we have revealed that simple density-based ITA functions are in good relationships with molecular polarizabilities for a series of 30 hydrogen-bonded base pairs and all 20 natural α-amino acids, 400 dipeptides, and 8000 tripeptides. Based on these lines, one can easily predict the molecular polarizabilities of larger peptides, even proteins as long as the total molecular wavefunction is available, rather than solving the computationally demanding coupled-perturbed Hartree-Fock (CPHF) equation or its DFT counterpart coupled-perturbed Kohn-Sham (CPKS) equation.

Spondyloarthritis comprises of a group of inflammatory diseases of the joints and spine with various clinical manifestations. The group includes ankylosing spondylitis, reactive arthritis, psoriatic arthritis, arthritis associated with inflammatory bowel disease, and undifferentiated spondyloarthritis. The exact etiology and pathogenesis of spondyloarthritis are still unknown, but five hypotheses explaining the pathogenesis exist. These hypotheses suggest that spondyloarthritis is caused by arthritogenic peptides, an unfolded protein response, HLA-B27 homodimer formation, malfunctioning endoplasmic reticulum aminopeptidases, and, last but not least, gut inflammation and dysbiosis. Here we discuss the five hypotheses and the evidence supporting each. In all of these hypotheses, HLA-B27 plays a central role. It is likely that a combination of these hypotheses, with HLA-B27 taking center stage, will eventually explain the development of spondyloarthritis in predisposed individuals.

Even though Alzheimer’s disease (AD) is of significant interest to the scientific community, its pathogenesis is very complicated and not well-understood. A great deal of progress has been made in AD research recently and with the advent of these new insights more therapeutic benefits may be identified that could help patients around the world. Much of the research in AD thus far has been very neuron-oriented; however, recent studies suggest that glial cells, i.e., microglia, astrocytes, oligodendrocytes, and oligodendrocyte progenitor cells (NG2 glia), are linked to the pathogenesis of AD and may offer several potential therapeutic targets against AD. In addition to a number of other functions, glial cells are responsible for maintaining homeostasis (i.e., concentration of ions, neurotransmitters, etc.) within the central nervous system (CNS) and are crucial to the structural integrity of neurons. This review explores the: (i) role of glial cells in AD pathogenesis; (ii) complex functionalities of the components involved; and (iii) potential therapeutic targets that could eventually lead to a better quality of life for AD patients

Recent studies have elucidated the diversity of genes encoding venom in sea anemones. However, most of those genes are yet to be explored in an evolutionary context. Insulin is a common peptide across metazoans and has been coopted into a predatory venom in many venomous lineages. In this study, we focus on the diversity of insulin-derived venoms in sea anemones and on elucidating their evolutionary history. We sourced data for 34 species of sea anemones and found sequences belonging to two venom families that have Insulin PFAM annotations (Cono-Insulin, VP302). Our findings show that both families have undergone duplication events. Members of each of the independently evolving clades have consistent predicted protein structures and distinct dN/dS values. Our work also shows that VP302 is part of a multidomain venom contig and has experienced a secondary gain into the venom system of cuticulate sea anemones.

The urgent global health challenge posed by methicillin-resistant Staphylococcus aureus (MRSA) infections demands effective solutions. Antimicrobial peptides (AMPs) represent promising tools of research of new antibacterial agents and LyeTx I mn∆K, a short synthetic peptide based on the Lycosa erythrognatha spider venom, is a good representative. This study focused on analyzing some antimicrobial activities of LyeTx I mn∆K, including minimum inhibitory and bactericidal concentrations, synergy and resensitization assays, lysis activity, the effect on biofilm, and the bacterial death curve in MRSA. Additionally, its characterization was conducted through isothermal titration calorimetry, dynamic light scattering, calcein release, and finally, efficacy in a mice wound model. The peptide demonstrating remarkable efficacy against planktonic cells and biofilms of MRSA, outperforms vancomycin in terms of rapid bactericidal action and anti-biofilm effects. The mechanism involves significant membrane damage. Interactions with bacterial model membranes, including those with lysylphosphatidylglycerol (LysylPOPG) modifications, highlight versatility and selectivity of this compound. Also, the peptide has the ability to sensitize resistant bacteria to conventional antibiotics, showing potential for combinatory therapy. Furthermore, using an in vivo model, was showed that a formulated gel containing the peptide proved superior to vancomycin in treating MRSA-induced wound in mice. Together the results highlight LyeTx I mnΔK as a promising prototype for the development of effective therapeutic strategies against superficial MRSA infections.

Phosphodiesterases (PDEs) are the principal superfamily of enzymes responsible for degrading the secondary messengers 3’,5’-cyclic nucleotides cAMP and cGMP. Their refined subcellular localization and substrate specificity contribute to finely regulate cAMP/cGMP gradients in various cellular microdomains. Redistribution of multiple signal compartmentalization components is often perceived under pathological conditions. Thereby PDEs have long been pursued as therapeutic targets in diverse disease conditions including neurological, metabolic, cancer and autoimmune disorders in addition to numerous cardiovascular diseases. PDE2 is a unique member of the broad family of PDEs. In addition to its capability to hydrolyze both cAMP and cGMP, PDE2 is the sole isoform that may be allosterically activated by cGMP increasing its cAMP hydrolyzing activity. Within the cardiovascular system, PDE2 serves as an integral regulator for the crosstalk between cAMP/cGMP pathways and thereby may couple chronically adverse augmented cAMP signalling with cardioprotective cGMP signalling. This review provides a comprehensive overview of PDE2 regulatory functions in multiple cellular components within the cardiovascular system and also within various subcellular microdomains. Implications for PDE2 mediated crosstalk mechanisms in diverse cardiovascular pathologies are discussed highlighting the prospective use of PDE2 as a potential therapeutic target in cardiovascular disorders.

Background: There are no known medicines or vaccines to control the COVID-19 pandemic caused by SARS-CoV-2 (nCoV). Antiviral peptides are superior to conventional drugs and may also be effective against COVID-19. Hence, we investigated the SARS-CoV-2 Spike RBD (nCoV-RBD) that interacts with hACE2 for viral attachment and entry. Methods: Three strategies and bioinformatics approaches were employed to design potential nCoV-RBD - hACE2 interaction-blocking peptides that may restrict viral attachment and entry. Firstly, the key residues interacting with nCoV-RBD - hACE2 are identified and hACE2 sequence based peptides are designed. Second, peptides from five antibacterial peptide databases that block nCoV-RBD are identified; finally, a chimeric peptide design approach is used to design peptides that can bind to key nCoV-RBD residues. The final peptides are selected based on their physiochemical properties, numbers and positions of key residues binding, binding energy, and antiviral properties. Results: We found (i) three amino acid stretches in hACE2 interact with nCoV-RBD; (ii) effective peptides must bind to three key positions of nCoV-RBD: Gly485/Phe486/Asn487, Gln493, and Gln498/Thr500/Asn501; (iii) Phe486, Gln493, and Asn501 are critical residues; (iv) AC20 and AC23 derived from hACE2 may block two key critical positions; (iv) DBP6 identified from databases can block the three sites of the nCoV-RBD interacting with one critical position Gln498; (v) seven chimeric peptides were considered promising among which cnCoVP-3, cnCoVP-4, and cnCoVP-7 are the top three; and (vi) cnCoVP-4 meets all the criteria and is the best peptide. Conclusion: All the ten peptides need experimental validation for their therapeutic efficacy.