The rise in antimicrobial resistance represents a significant challenge to global health. The reason partially lies in an inappropriate use of conventional antibiotics and the subsequent rapid spread of multidrug-resistant pathogen strains. This emergency requires an urgent search for conceptually new antimicrobial agents. A viable alternative to conventional antibiotics is antimicrobial peptides (AMPs), which are ribosomally synthesized molecules with considerable potential as next-generation anti-infectious therapeutics. Previously, we have reported on the β-hairpin peptide Ap9, an analog of abarenicin from the marine polychaeta Abarenicola pacifica, with potent activity against key Gram-negative pathogens. Here, it is shown that Ap9 acts in a manner resembling polymyxin B, namely via interaction with lipopolysaccharide (LPS), and retains its activity against polymyxin-resistant isolates without observed cross-resistance, and causes insignificant damage in cytoplasmic membrane at bactericidal concentrations. NMR spectroscopy reveals that LPS binding induces a conformational rearrangement of Ap9, its dimer formation, and local structural remodeling of the peptide region (residues 8-12) into 310-helix. Bacterial resistance to Ap9 was found to be relatively low with a reduced susceptibility associated with infrequent genetic alterations, such as the mutation in lptD or the deletion in mlaA. Furthermore, Ap9 demonstrates a favorable tolerability, a wider therapeutic window than that of polymyxin B, and a sufficiently long half-life through the systemic use, as well as in vivo efficacy in murine models of Gram-negative infections, including sepsis caused by the mcr-1-harboring Escherichia coli strain. The obtained results point to Ap9 as a promising candidate for further preclinical studies aimed at development of an alternative to polymyxins.

Liposomal systems are commonly used in drug delivery due to their low toxicity, biocompatibility and biodegradability. In this review, the influence of physicochemical parameters of liposomes, namely size, surface charge, and lipid composition, on biodistribution, dendritic cell uptake, and the character of the induced immune response is examined. The potential of strategies such as targeting C-type lectin receptors, the combined use of toll-like receptor agonists and tolerogenic molecules, and an approach based on high-affinity antigen binding to liposomes via coiled coil-forming peptides is evaluated. Mechanisms of tolerance induction at the humoral, cytokine, and cellular levels are discussed, including the switch from a Th2 to a regulatory T-cell response and the formation of blocking antibodies. Special attention is paid to safety aspects associated with the use of liposomal systems, specifically avoidance of pseudoallergic reactions linked to the complement system activation and the toxicity of cationic lipids, as well as approaches for minimization of these risks. The main obstacles to clinical application and promising directions of further research necessary for the development of effective and safe liposomal allergy vaccines are outlined. This review summarizes current data on the use of liposomal systems for allergen-specific immunotherapy.

The skin serves as the first line barrier of innate immunity, protecting the body from external influences and maintaining its homeostasis. Exogenous and endogenous stress factors alter the structure and functional properties of the skin. The search for compounds capable of counteracting these processes has allowed the identification of peptides as promising ingredients of products for medicinal and cosmetic applications. This review comprehensively examines the mechanisms of action and dermatological applications of two distinct classes of natural products—endogenous human peptides and those derived from marine organisms. Human peptides exhibit numerous biological functions, including antimicrobial and immunomodulatory ones, as well as promoting antioxidant protection and wound healing. Microbiome-associated peptides are an underestimated but powerful regulator of skin aging through immunomodulation, inflammation control, barrier function maintenance, and selection of the proper microbial community. Peptides from marine organisms exhibit significant structural diversity and a broad spectrum of biological activity, including regenerative effects and effects on antibiotic-resistant microorganisms. This review summarizes current data obtained from in vitro, ex vivo, and clinical studies demonstrating a broad potential of peptides for maintaining skin health. Both peptide classes represent powerful, targeted strategies for innovative dermatological interventions aimed at promoting skin rejuvenation, protection, and overall homeostasis.

Background/Objectives: Currently, infections caused by fungi of the Candida genus remain a significant global health concern. The rising incidence of mycoses, coupled with the rapid emergence of fungal resistance, highlights the urgent need to search for new antifungal agents. Here, we obtained the recombinant hevein-like peptide from Amaranthus caudatus with two amino acid substitutions (F18W in the chitin-binding motif and M13A preventing the peptide from cleavage with cyanogen bromide during its biotechnological production). Methods: Antifungal potential of the modified hevein-like peptide, designated as mAc-AMP2, against susceptible and resistant strains of Candida albicans and non-albicans Candida species was studied. Results: We showed that mAc-AMP2 possessed anticandidal activities against all strains tested at nanomolar peptide concentrations. The presence of salts or serum affected the action of the peptide but its antifungal activity remained quite high. mAc-AMP2 exhibited anti-adherent properties and inhibited the formation of fungal biofilms. Using RP-HPLC, we demonstrated that degradation of the peptide in the presence of serum occurred rather slowly. mAc-AMP2 did not exhibit hemolytic and cytotoxic activities against the Caco-2 cell monolayer and peripheral blood mononuclear cells. Using flow cytometry, we demonstrated that the peptide at its high concentrations increased fungal membrane permeability. In resistance induction experiments, sensitivity of C. albicans toward mAc-AMP2 decreased over time, but restored after the peptide elimination. Conclusions: Taking into account all the data obtained, we suggest that the modified hevein-like peptide is a promising candidate for development of novel therapeutic agents to combat fungal infections caused by C. albicans and other Candida species.

The birch pollen allergen Bet v 1 is believed to be the main sensitizer among PR-10 allergens. Recent data have shown that some other PR-10 allergens also display sensitization activities, and Bet v 1-based immunotherapy is not effective for blocking allergic reactions to PR-10 proteins with low similarities to Bet v 1. Here, we investigated the sensitization potential of the major soybean allergen Gly m 4 and its cross-reactivity with Bet v 1. We demonstrated that Gly m 4 bound cholesterol and bile acids, including deoxycholic acid (DCA). Using qPCR, we showed that Gly m 4 induced the expression of genes encoding alarmins TSLP and IL-33 in intestinal-like Caco-2 cells; however, its fragments resulting from digestion by gastroduodenal enzymes or the DCA-bound Gly m 4 caused more pronounced gene upregulation. Using competitive ELISA, we demonstrated the low cross-reactivity of anti-Gly m 4 and anti-Bet v 1 IgG, raised in laboratory animals. Using mice allergy models with sensitization to birch or soybean allergens, we also showed a low cross-reactivity of Gly m 4- and Bet v 1-specific IgE, IgG1 and IgG2a. Thus, our findings support an assumption of the intrinsic sensitization capacity of Gly m 4 and the existence of Gly m 4-specific antibodies in sera of allergic patients.

Gly m 4, a soybean PR-10 allergen, is known to trigger systemic allergic reactions. However, the intrinsic sensitizing potential of the allergen remains unclear. Adjuvant-free murine models of sensitization to Gly m 4 might help to investigate mechanisms of a soy allergy and establish relevant in vivo platforms for developing novel allergen-specific immunotherapy strategies. BALB/c mice were sensitized to Gly m 4 via intraperitoneal (i.p.), subcutaneous (s.c.), or intranasal (i.n.) routes, with or without adjuvant (alum or lipopolysaccharide (LPS)). In order to assess sensitization, we evaluated levels of allergen-specific IgE, IgG1, IgG2a, systemic anaphylaxis, rat basophil (RBL) degranulation, and cytokine/chemokine profiles in mouse sera. I.n. exposure with or without LPS proved to be ineffective and did not elicit sensitization. I.p. and s.c. routes of sensitization with and without alum induced a Th2-skewed response, which was demonstrated by high levels of IgE and IgG1, systemic anaphylaxis, and IgE-mediated degranulation of RBL cells. Adjuvant-free i.p. administration led to a shift in cytokine production, with reduced levels of proinflammatory (IL-1α/IL-6) cytokines and increased levels of Th2-associated (IL-13/GM-CSF) ones. Thus, adjuvant-free murine models validated the intrinsic sensitizing capacity of Gly m 4. Moreover, Gly m 4 demonstrated similar immunogenic profiles to Bet v 1 in alum-based models. It is the first evidence that soybean Gly m 4 can induce in vivo allergic sensitization in mice without adjuvants, particularly via i.p. and s.c. routes. Established adjuvant-free murine models offer a relevant tool for studying soy allergy and developing targeted immunotherapy.

Marine peptides, derived from a great number of aquatic organisms, exhibit a broad spectrum of biological activities that hold a significant therapeutic potential. This article reviews the multifaceted roles of marine peptides, focusing on their antibacterial, antibiofilm, antifungal, antiviral, antiparasitic, cytotoxic, anticancer, immunomodulatory, chemotactic, opsonizing, anti-inflammatory, antiaging, skin-protective, and wound-healing properties. By elucidating mechanisms of their action and highlighting key research findings, this review aims to provide a comprehensive understanding of possible therapeutic applications of marine peptides, underscoring their importance in developing novel drugs as well as in cosmetology, food industry, aquatic and agriculture biotechnology. Further investigations are essential to harness their therapeutic potential and should focus on detailed mechanism studies, large-scale production, and clinical evaluations with a view to confirm their efficacy and safety and translate these findings into practical applications. It is also important to investigate the potential synergistic effects of marine peptide combinations with existing medicines to enhance their efficacy. Challenges include the sustainable sourcing of marine peptides, and therefore an environmental impact of harvesting marine organisms must be considered as well.

Pollen allergy is rated as a major public health problem, causing significant morbidity and adversely affecting the quality of people’s lives. The airway epithelium serves as the first line of defense in the respiratory system, playing a crucial role in orchestrating immune responses to allergens. In this work, we studied the important transport steps in the major alder pollen allergen Aln g 1 through the human airway epithelium in comparison with those of the birch pollen allergen Bet v 1. Using fluorescence spectroscopy, we showed that both allergens can destroy liposomes with a composition modeling the adult human pulmonary surfactant. Using a polarized Calu-3 monolayer, we showed similar efficiencies of Aln g 1 and Bet v 1 transport through the artificial epithelial barrier. Using qPCR, we showed that Aln g 1 upregulates the expression of IL-33, TSLP, IL-1β, CXCL8 in epithelial cells, playing an important role in the sensitization process. The obtained results may improve our understanding of the primary sensitization mechanisms with the involvement of the PR-10 family of lipid-binding allergens.

Background/Objectives: The growing resistance development among fungi, including those of Candida species, poses significant challenges to public health, emphasizing the need for the implementation of innovative therapeutic approaches. The tobacco defensin NaD1 exhibits a pronounced activity against C. albicans, but its relatively high cytotoxicity toward mammalian cells limits its potential application. Here, we investigated anticandidal activity and cytotoxicity of four modified analogues of NaD1 (NaD1-1 T44R/K45R, NaD1-2 L38R, NaD1-3 K36R/L38R, NaD1-4 L38R/T44R/K45R). Methods: These peptides contained substitutions with arginine of some amino acid residues in the C-terminal region of NaD1 and in its L5 loop (S35KILRR40), responsible for the “cationic grip” and binding to phosphatidylinositol 4,5-bisphosphate (PIP4,5), one of the primary targets of tobacco defensin action. Results: We showed that the modified NaD1 analogues effectively inhibited the growth of C. albicans cells but had a less fungicidal action than NaD1. As compared to NaD1, its modified analogues differed in their sensitivity to the presence of various salts; antifungal activities of NaD1-3 and NaD1-4 were more tolerant to the presence of NaCl and CaCl2, respectively. All modified analogues except NaD1-1 did not exhibit hemolytic activity and showed significantly less cytotoxicity towards human immune and epithelial cells compared to NaD1. All modified analogues enhanced the permeability of PIP4,5-containing liposomes, although less effectively than NaD1. Differences in their properties were also demonstrated through experiments on oligomerization and zymosan binding. Conclusions: Thus, we proposed that the modified NaD1 analogues NaD1-2, NaD1-3, and NaD1-4 appear to be promising candidate antifungals. However, further in vitro and in vivo studies are required to evaluate their therapeutic potential against critical fungal pathogens.

Currently, the spread of fungal infections is becoming an urgent problem. Fungi of the Candida genus are opportunistic microorganisms that cause superficial and life-threatening systemic candidiasis in immunocompromised patients. The list of antifungal drugs for the treatment of candidiasis is very limited, while the prevalence of resistant strains is growing rapidly. Therefore, the search for new antimycotics, including those exhibiting immunomodulatory properties, is of great importance. Plenty of natural compounds with antifungal activities may be extremely useful in solving this problem. This review evaluates the features of natural antimicrobial peptides, namely plant defensins as possible prototypes of new anticandidal agents. Plant defensins are important components of the innate immune system, which provides the first line of defense against pathogens. The introduction presents a brief summary regarding pathogenic Candida species, the pathogenesis of candidiasis, and the mechanisms of antimycotic resistance. Then, the structural features of plant defensins, their anticandidal activities, their mechanisms of action on yeast-like fungi, their ability to prevent adhesion and biofilm formation, and their combined action with conventional antimycotics are described. The possible mechanisms of fungal resistance to plant defensins, their cytotoxic activity, and their effectiveness in in vivo experiments are also discussed. In addition, for the first time for plant defensins, knowledge about their immunomodulatory effects is also presented.

The adaption of plants to stressful environments depends on long-distance responses in plant organs, which themselves are remote from sites of perception of external stimuli. Jasmonic acid (JA) and its derivatives are known to be involved in plants’ adaptation to salinity. However, to our knowledge, the transport of JAs from roots to shoots has not been studied in relation to the responses of shoots to root salt treatment. We detected a salt-induced increase in the content of JAs in the roots, xylem sap, and leaves of pea plants related to changes in transpiration. Similarities between the localization of JA and lipid transfer proteins (LTPs) around vascular tissues were detected with immunohistochemistry, while immunoblotting revealed the presence of LTPs in the xylem sap of pea plants and its increase with salinity. Furthermore, we compared the effects of exogenous MeJA and salt treatment on the accumulation of JAs in leaves and their impact on transpiration. Our results indicate that salt-induced changes in JA concentrations in roots and xylem sap are the source of accumulation of these hormones in leaves leading to associated changes in transpiration. Furthermore, they suggest the possible involvement of LTPs in the loading/unloading of JAs into/from the xylem and its xylem transport.

Today, allergies have become a serious problem. PR-10 proteins are clinically relevant allergens that have the ability to bind hydrophobic ligands, which can significantly increase their allergenicity potential. It has been recently shown that not only the birch pollen allergen Bet v 1 but also the alder pollen allergen Aln g 1, might act as a true sensitizer of the immune system. The current investigation is aimed at the further study of the allergenic and structural features of Aln g 1. By using qPCR, we showed that Aln g 1 was able to upregulate alarmins in epithelial cells, playing an important role in sensitization. With the use of CD-spectroscopy and ELISA assays with the sera of allergic patients, we demonstrated that Aln g 1 did not completely restore its structure after thermal denaturation, which led to a decrease in its IgE-binding capacity. Using site-directed mutagenesis, we revealed that the replacement of two residues (Asp27 and Leu30) in the structure of Aln g 1 led to a decrease in its ability to bind to both IgE from sera of allergic patients and lipid ligands. The obtained data open a prospect for the development of hypoallergenic variants of the major alder allergen Aln g 1 for allergen-specific immunotherapy.

A total chemical synthesis of spacer-armed Forssman pentasaccharide is reported. The choice of the 2(donor) + 3(acceptor) block scheme, the optimal combination of a limited number of simple protecting groups and the sequence of deprotection steps allowed to achieve the high yield and stereoselectivity of glycosylation and to avoid losses during deprotection. The target pentasaccharide was obtained in a 10-mg scale. 1H and 13C NMR spectra of the Forssman pentasaccharide were completely assigned with the use of various 2D-NMR experiments.

Background: Antimicrobial peptides are generally considered promising drug candidates for combating resistant bacterial infections. However, the selectivity of their action may vary significantly. Natural gomesin, isolated from haemocytes of the tarantula Acanthoscurria gomesiana, demonstrates a broad spectrum of antimicrobial activities, being the most effective against pathogenic fungi. Methods: Here, we searched for variants of natural gomesin-like peptides and produced their recombinant analogs in the bacterial expression system. The antimicrobial activities of the obtained peptides were tested against a panel of bacterial and yeast strains, and their toxicity towards human cells was examined. Results: Most of the new analogs of gomesin have primary structures homologous to that of the natural gomesin; however, they have fewer amino acid residues and post-translational modifications. One of the discovered analogs, the His-rich shorter peptide from the spider Dysdera sylvatica, designated as DsGom, displays antifungal activity comparable with that of natural gomesin. In the process of the structural–functional study of DsGom, it was shown that this analog retains a basic mechanism of action similar to that of natural gomesin. The DsGom analog has a significantly better toxicity profile as compared to gomesin. At the same time, the loss of the first Arg residue reduces, but does not annul, the antifungal activity of DsGom. Moreover, the acidification of the growth medium reduces the loss of the antifungal activity of this analog. Conclusions: The discovered natural gomesin-like peptides display more selective antifungal activities as compared to gomesin. The low cytotoxicity of DsGom, combined with its high antifungal activity and stability, allows us to consider it a promising drug candidate for the treatment of fungal infections, especially those caused by fungi of the Candida genus.

Adaptation to changes in the environment depends, in part, on signaling between plant organs to integrate adaptive response at the level of the whole organism. Changes in the delivery of hormones from one organ to another through the vascular system strongly suggest that hormone transport is involved in the transmission of signals over long distances. However, there is evidence that, alternatively, systemic responses may be brought about by other kinds of signals (e.g., hydraulic or electrical) capable of inducing changes in hormone metabolism in distant organs. Long-distance transport of hormones is therefore a matter of debate. This review summarizes arguments for and against the involvement of the long-distance transport of cytokinins in signaling mineral nutrient availability from roots to the shoot. It also assesses the evidence for the role of abscisic acid (ABA) and jasmonates in long-distance signaling of water deficiency and the possibility that Lipid-Binding and Transfer Proteins (LBTPs) facilitate the long-distance transport of hormones. It is assumed that proteins of this type raise the solubility of hydrophobic substances such as ABA and jasmonates in hydrophilic spaces, thereby enabling their movement in solution throughout the plant. This review collates evidence that LBTPs bind to cytokinins, ABA, and jasmonates and that cytokinins, ABA, and LBTPs are present in xylem and phloem sap and co-localize at sites of loading into vascular tissues and at sites of unloading from the phloem. The available evidence indicates a functional interaction between LBTPs and these hormones.

The synthesis of glycoprobes based on water-soluble polyacrylamide with a custom content of any low molecular weight fluorescent label is described. The synthesis is carried out in a stepwise manner: in the first step, the polymer is strictly dose-modified with a side substituent-linker containing an amino group; in the second step, the amino group is quantitatively acylated with an activated ester of the dye. The probes are designed to identify and study carbohydrate-binding proteins of plant cells.

Today, Candida albicans is still the most common cause of both local and life-threatening systemic candidiasis. The spread of resistant fungal strains has resulted in an urgent need to search for new promising antimycotics. Here, we investigated the antifungal action of the tobacco defensin NaD1 against susceptible and resistant to azoles and echinocandins strains of C. albicans. We demonstrated that NaD1 was equally effective and fungicidal against all tested strains. The MIC and MFC values were 6.25 and 12.5 µM, respectively. We showed for the first time that NaD1 could act synergistically not only with caspofungin but also with human host defense antimicrobial peptides cathelicidin LL-37 and β-defensin-2 (HBD2) against susceptible and resistant fungal strains. Using flow cytometry, we demonstrated that NaD1 in combinations with LL-37 or HBD2 can reinforce each other by enhancing membrane disruption. Using the Caco-2 cell monolayer model, we demonstrated that NaD1 impaired the adhesion of C. albicans cells to the human epithelium. Moreover, NaD1 inhibited the formation of fungal biofilms in Sabouraud broth and less markedly in nutrient-rich RPMI-1640 medium, and enhanced the antibiofilm activity of caspofungin. Thus, we hypothesized that NaD1 might affect the development of candidiasis in vivo, including that caused by resistant fungal strains.

Polysaccharides are widely represented in plants and bacteria, where they are distinguished by their extraordinary structural diversity and the performance of various functions. To study the functions of polysaccharides, their fluorescent labeling is needed. This review discusses methods for introducing fluorescent labels into polysaccharides by chemical modification of certain functional groups of these complex biopolymers, as well as using the so-called bioorthogonal reactions, which allow labeling in a cell without affecting its viability. In addition to modification with organic dyes, the possibility of using quantum dots and coordination compounds of lanthanides is also discussed.

Capitellacin is the β-hairpin membrane-active cationic antimicrobial peptide from the marine polychaeta Capitella teleta. Capitellacin exhibits antibacterial activity, including against drug-resistant strains. To gain insight into the mechanism of capitellacin action, we investigated the structure of the peptide in the membrane-mimicking environment of dodecylphosphocholine (DPC) micelles using high-resolution NMR spectroscopy. In DPC solution, two structural forms of capitellacin were observed: a monomeric β-hairpin was in equilibrium with a dimer formed by the antiparallel association of the N-terminal β-strands and stabilized by intermonomer hydrogen bonds and Van der Waals interactions. The thermodynamics of the enthalpy-driven dimerization process was studied by varying the temperature and molar ratios of the peptide to detergent. Cooling the peptide/detergent system promoted capitellacin dimerization. Paramagnetic relaxation enhancement induced by lipid-soluble 12-doxylstearate showed that monomeric and dimeric capitellacin interacted with the surface of the micelle and did not penetrate into the micelle interior, which is consistent with the “carpet” mode of membrane activity. An analysis of the known structures of β-hairpin AMP dimers showed that their dimerization in a membrane-like environment occurs through the association of polar or weakly hydrophobic surfaces. A comparative analysis of the physicochemical properties of β-hairpin AMPs revealed that dimer stability and hemolytic activity are positively correlated with surface hydrophobicity. An additional positive correlation was observed between hemolytic activity and AMP charge. The data obtained allowed for the provision of a more accurate description of the mechanism of the oligomerization of β-structural peptides in biological membranes.

Within class II bacteriocins, we assume the presence of a separate subfamily of antimicrobial peptides possessing a broad spectrum of antimicrobial activity. Although these peptides are structurally related to the subclass IIa (pediocin-like) bacteriocins, they have significant differences in biological activities and, probably, a mechanism of their antimicrobial action. A representative of this subfamily is acidocin A from Lactobacillus acidophilus TK9201. We discovered the similarity between acidocin A and acidocin 8912 from Lactobacillus acidophilus TK8912 when analyzing plasmids from lactic acid bacteria and suggested the presence of a single evolutionary predecessor of these peptides. We obtained the C-terminally extended homolog of acidocin 8912, named acidocin 8912A, a possible intermediate form in the evolution of the former. The study of secondary structures and biological activities of these peptides showed their structural similarity to acidocin A; however, the antimicrobial activities of acidocin 8912 and acidocin 8912A were lower than that of acidocin A. In addition, these peptides demonstrated stronger cytotoxic and membranotropic effects. Building upon what we previously discovered about the immunomodulatory properties of acidocin A, we studied its proteolytic stability under conditions simulating those in the digestive tract and also assessed its ability to permeate intestinal epithelium using the Caco-2 cells monolayer model. In addition, we found a pronounced effect of acidocin A against fungi of the genus Candida, which might also expand the therapeutic potential of this bacterial antimicrobial peptide.