Search Results (137)

Leucine residues are commonly found in the hydrophobic face of antimicrobial peptides (AMPs) and are crucial for membrane permeabilization, leading to the cell death of invading pathogens. Melittin, which contains four leucine residues, demonstrates broad-spectrum antimicrobial properties but also significant cytotoxicity against mammalian cells. To enhance the cell selectivity of melittin, this study synthesized five analogs by replacing leucine with its structural isomer, 6-aminohexanoic acid. Among these analogs, Mel-LX3 exhibited potent antibacterial activity against both Gram-positive and Gram-negative bacteria. Importantly, Mel-LX3 displayed significantly reduced hemolytic and cytotoxic effects compared to melittin. Mechanistic studies, including membrane depolarization, SYTOX green uptake, FACScan analysis, and inner/outer membrane permeation assays, demonstrated that Mel-LX3 effectively permeabilized bacterial membranes similar to melittin. Notably, Mel-LX3 showed robust antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA) and multidrug-resistant Pseudomonas aeruginosa (MDRPA). Furthermore, Mel-LX3 effectively inhibited biofilm formation and eradicated existing biofilms of MDRPA. With its improved selective antimicrobial and antibiofilm activities, Mel-LX3 emerges as a promising candidate for the development of novel antimicrobial agents. We propose that the substitution of leucine with 6-aminohexanoic acid in AMPs represents a significant strategy for combating resistant bacteria. Full article

Antimicrobial peptides (AMPs) are being explored as a potential strategy to combat antibiotic resistance due to their ability to reduce susceptibility to antibiotics. This study explored whether the [R₄W₄] peptide mode of action is bacteriostatic or bactericidal using modified two-fold serial dilution and evaluating the synergism between gentamicin and [R₄W₄] against Escherichia coli (E. coli) and methicillin-resistant Staphylococcus aureus (MRSA) by a checkered board assay. [R₄W₄] exhibited bactericidal activity against bacterial isolates (MBC/MIC ≤ 4), with a synergistic effect with gentamicin against E. coli (FICI = 0.3) but not against MRSA (FICI = 0.75). Moreover, we investigated the mechanism of action of [R₄W₄] against MRSA by applying biophysical assays to evaluate zeta potential, cytoplasmic membrane depolarization, and lipoteichoic acid (LTA) binding affinity. [R₄W₄] at a 16 mg/mL concentration stabilized the zeta potential of MRSA −31 ± 0.88 mV to −8.37 mV. Also, [R₄W₄] at 2 × MIC and 16 × MIC revealed a membrane perturbation process associated with concentration-dependent effects. Lastly, in the presence of BODIPY-TR-cadaverine (BC) fluorescence dyes, [R₄W₄] exhibited binding affinity to LTA comparable with melittin, the positive control. In addition, the antibacterial activity of [R₄W₄] against MRSA remained unchanged in the absence and presence of LTA, with an MIC of 8 µg/mL. Therefore, the [R₄W₄] mechanism of action is deemed bactericidal, involving interaction with bacterial cell membranes, causing concentration-dependent membrane perturbation. Additionally, after 30 serial passages, there was a modest increment of MRSA strains resistant to [R₄W₄] and a change in antibacterial effectiveness MIC [R₄W₄] and vancomycin by 8 and 4 folds with a slight change in Levofloxacin MIC 1 to 2 µg/mL. These data suggest that [R₄W₄] warrants further consideration as a potential AMP. Full article

Animal venoms are intricate and teem with potential for groundbreaking medical advancements. Although traditional methods for purifying venom proteins are effective, they usually require complicated, multi-step processes that lead to lower yields. Our study introduces an efficient, one-step technique for extracting venom-derived proteins through reverse-phase high-performance liquid chromatography (RP-HPLC). We carefully optimized the RP-HPLC process, focusing on the gradient elution conditions and the strategic use of our columns’ stationary phase characteristics, to enhance the effectiveness of our separations. This enabled us to efficiently isolate six venom proteins: melittin (2.846 kDa) from Apis mellifera with a yield of 4.5% and homogeneity of 99%; α-cobratoxin (7.821 kDa) from Naja kaouthia with a yield of 15% and homogeneity of 99%; α-bungarotoxin (7.983 kDa) from Bungarus multicinctus with a yield of 7% and purity of 99%; calciseptine (7.035 kDa) from Dendroaspis polylepis with a yield of 6% and homogeneity of 95%; notexin (13.593 kDa) from Notechis scutatus with a yield of 10% and homogeneity of 95%; and CVFm (150 kDa) from Naja melanoleuca with a yield of 0.8% and homogeneity of 94%. These were all accomplished in one step. This breakthrough simplifies the purification of venom peptides and proteins, making the process more feasible and economical. It paves the way for developing new drugs and promising treatments that are both more effective and precisely targeted. Full article

Beekeeping provides products with nutraceutical and pharmaceutical characteristics. These products are characterized by abundance of bioactive compounds. For different reasons, honey, royal jelly, propolis, venom, and pollen are beneficial to humans and animals and could be used as therapeutics. The pharmacological action of these products is related to many of their constituents. The main bioactive components of honey include oligosaccharides, methylglyoxal, royal jelly proteins (MRJPs), and phenolics compounds. Royal jelly contains jelleins, royalisin peptides, MRJPs, and derivatives of hydroxy-decenoic acid, particularly 10-hydroxy-2-decenoic acid (10-HDA), which possess antibacterial, anti-inflammatory, immunomodulatory, neuromodulatory, metabolic syndrome-preventing, and anti-aging properties. Propolis has a plethora of activities that are referable to compounds such as caffeic acid phenethyl ester. Peptides found in bee venom include phospholipase A2, apamin, and melittin. In addition to being vitamin-rich, bee pollen also includes unsaturated fatty acids, sterols, and phenolics compounds that express antiatherosclerotic, antidiabetic, and anti-inflammatory properties. Therefore, the constituents of hive products are particular and different. All of these constituents have been investigated for their properties in numerous research studies. This review aims to provide a thorough screening of the bioactive chemicals found in honeybee products and their beneficial biological effects. The manuscript may provide impetus to the branch of unconventional medicine that goes by the name of apitherapy. Full article

This research aimed to explore the healing impacts of Melittin treatment on gastrocnemius muscle wasting caused by immobilization with a cast in rabbits. Twenty-four rabbits were randomly allocated to four groups. The procedures included different injections: 0.2 mL of normal saline to Group 1 (G1-NS); 4 μg/kg of Melittin to Group 2 (G2-4 μg/kg Melittin); 20 μg/kg of Melittin to Group 3 (G3-20 μg/kg Melittin); and 100 μg/kg of Melittin to Group 4 (G4-100 μg/kg Melittin). Ultrasound was used to guide the injections into the rabbits’ atrophied calf muscles following two weeks of immobilization via casting. Clinical measurements, including the length of the calf, the compound muscle action potential (CMAP) of the tibial nerve, and the gastrocnemius muscle thickness, were assessed. Additionally, cross-sectional slices of gastrocnemius muscle fibers were examined, and immunohistochemistry and Western blot analyses were performed following two weeks of therapy. The mean regenerative changes, as indicated by clinical parameters, in Group 4 were significantly more pronounced than in the other groups (p < 0.05). Furthermore, the cross-sectional area of the gastrocnemius muscle fibers and immunohistochemical indicators in Group 4 exceeded those in the remaining groups (p < 0.05). Western blot analysis also showed a more significant presence of anti-inflammatory and angiogenic cytokines in Group 4 compared to the others (p < 0.05). Melittin therapy at a higher dosage can more efficiently activate regeneration in atrophied gastrocnemius muscle compared to lower doses of Melittin or normal saline. Full article

Among the various natural compounds used in alternative and Oriental medicine, toxins isolated from different organisms have had their application for many years, and Apis mellifera venom has been studied the most extensively. Numerous studies dealing with the positive assets of bee venom (BV) indicated its beneficial properties. The usage of bee products to prevent the occurrence of diseases and for their treatment is often referred to as apitherapy and is based mainly on the experience of the traditional system of medical practice in diverse ethnic communities. Today, a large number of studies are focused on the antitumor effects of BV, which are mainly attributed to its basic polypeptide melittin (MEL). Previous studies have indicated that BV and its major constituent MEL cause a strong toxic effect on different cancer cells, such as liver, lung, bladder, kidney, prostate, breast, and leukemia cells, while a less pronounced effect was observed in normal non-target cells. Their proposed mechanisms of action, such as the effect on proliferation and growth inhibition, cell cycle alterations, and induction of cell death through several cancer cell death mechanisms, are associated with the activation of phospholipase A₂ (PLA₂), caspases, and matrix metalloproteinases that destroy cancer cells. Numerous cellular effects of BV and MEL need to be elucidated on the molecular level, while the key issue has to do with the trigger of the apoptotic cascade. Apoptosis could be either a consequence of the plasmatic membrane fenestration or the result of the direct interaction of the BV components with pro-apoptotic and anti-apoptotic factors. The interaction of BV peptides and enzymes with the plasma membrane is a crucial step in the whole process. However, before its possible application as a remedy, it is crucial to identify the correct route of exposure and dosage of BV and MEL for potential therapeutic use as well as potential side effects on normal cells and tissues to avoid any possible adverse event. Full article

Herpes simplex virus (HSV) is known to cause cold sores and various diseases in humans. Importantly, HSV infection can develop latent and recurrent infections, and it is also known to cause inflammation. These infections are difficult to control, and effective treatment of the disease remains a challenge. Thus, the search for new antiviral and anti-inflammatory agents is a necessity. Melittin is a major peptide that is present in the venom of the honeybee. It possesses a number of pharmacological properties. In this study, the effects of the melittin peptides from A. mellifera (MEL-AM) and A. florea (MEL-AF) against HSV-1 and HSV-2 were evaluated at different stages during the viral multiplication cycle in an attempt to define the mode of antiviral action using plaque reduction and virucidal assays. The results revealed a new finding that melittin at 5 µg/mL demonstrated the highest inhibitory effect on HSV through the direct inactivation of viral particles, and MEL-AF displayed a greater virucidal activity. Moreover, melittin was also observed to interfere with the process of HSV attachment to the host cells. MEL-AM exhibited anti-HSV-1 and anti-HSV-2 effects with EC₅₀ values of 4.90 ± 0.15 and 4.39 ± 0.20 µg/mL, while MEL-AF demonstrated EC₅₀ values of 4.47 ± 0.21 and 3.95 ± 0.61 µg/mL against HSV-1 and HSV-2, respectively. However, non-cytotoxic concentrations of both types of melittin produced only slight degrees of HSV-1 and HSV-2 inhibition after viral attachment, but melittin at 5 µg/mL was able to reduce the plaque size of HSV-2 when compared to the untreated group. In addition, MEL-AM and MEL-AF also exhibited anti-inflammatory activity via the inhibition of nitric oxide production in LPS-stimulated RAW 264.7 macrophage cells, and they were also found to down-regulate the expressions of the iNOS, COX-2 and IL-6 genes. The highest inhibition of IL-6 mRNA expression was found after treatment with 10 µg/mL of MEL-AM and MEL-AF. Therefore, melittin peptides have displayed strong potential to be used as an alternative treatment for HSV infection and inflammatory diseases in the future. Full article

The development of antibiotic-resistant microorganisms is a major global health concern. Recently, there has been an increasing interest in antimicrobial peptides as a therapeutic option. This study aimed to evaluate the triple-action (broad-spectrum antibacterial, anti-biofilm, and anti-quorum sensing activities) of melittin, a membrane-active peptide present in bee venom. The minimum inhibitory concentration and minimum bactericidal concentration of the melittin were determined using the microdilution method and agar plate counting. Growth curve analysis revealed that melittin showed a concentration-dependent antibacterial activity. Scanning electron microscope analysis revealed that melittin treatment altered the morphology. Confocal laser scanning microscope revealed that melittin increased the membrane permeability and intracellular ROS generation in bacteria, all of which contribute to bacterial cell death. In addition, the crystal violet (CV) assay was used to test the anti-biofilm activity. The CV assay demonstrated that melittin inhibited biofilm formation and eradicated mature biofilms. Biofilm formation mediated by quorum sensing (QS) plays a major role in this regard, so molecular docking and molecular dynamics analysis confirmed that melittin interacts with LasR receptors through hydrogen bonds, and further evaluates the anti-QS activity of melittin through the production of virulence factors (pyocyanin, elastase, and rhamnolipid), exopolysaccharides secretion, and bacterial motility, that may be the key to inhibiting the biofilm formation mechanism. The present findings highlight the promising role of melittin as a broad-spectrum antibacterial, anti-biofilm agent, and potential QS inhibitor, providing a new perspective and theoretical basis for the development of alternative antibiotics. Full article

In recent years, there has been a growing interest in antimicrobial peptides as innovative antimicrobial agents for combating drug-resistant bacterial infections, particularly in the fields of biofilm control and eradication. In the present study, a novel cationic antimicrobial peptide, named LC-AMP-F1, was derived from the cDNA library of the Lycosa coelestis venom gland. The sequence, physicochemical properties and secondary structure of LC-AMP-F1 were predicted and studied. LC-AMP-F1 was tested for stability, cytotoxicity, drug resistance, antibacterial activity, and antibiofilm activity in vitro compared with melittin, a well-studied antimicrobial peptide. The findings indicated that LC-AMP-F1 exhibited inhibitory effects on the growth of various bacteria, including five strains of multidrug-resistant bacteria commonly found in clinical settings. Additionally, LC-AMP-F1 demonstrated effective inhibition of biofilm formation and disruption of mature biofilms. Furthermore, LC-AMP-F1 exhibited favorable stability, minimal hemolytic activity, and low toxicity towards different types of eukaryotic cells. Also, it was found that the combination of LC-AMP-F1 with conventional antibiotics exhibited either synergistic or additive therapeutic benefits. Concerning the antibacterial mechanism, scanning electron microscopy and SYTOX Green staining results showed that LC-AMP-F1 increased cell membrane permeability and swiftly disrupted bacterial cell membranes to exert its antibacterial effects. In summary, the findings and studies facilitated the development and clinical application of novel antimicrobial agents. Full article

Melittin, a natural antimicrobial peptide, has broad-spectrum antimicrobial activity. This has resulted in it gaining increasing attention as a potential antibiotic alternative; however, its practical use has been limited by its weak antimicrobial activity, high hemolytic activity, and low proteolytic stability. In this study, N-terminal fatty acid conjugation was used to develop new melittin-derived lipopeptides (MDLs) to improve the characteristics of melittin. Our results showed that compared with native melittin, the antimicrobial activity of MDLs was increased by 2 to 16 times, and the stability of these MDLs against trypsin and pepsin degradation was increased by 50 to 80%. However, the hemolytic activity of the MDLs decreased when the length of the carbon chain of fatty acids exceeded 10. Among the MDLs, the newly designed analog Mel-C8 showed optimal antimicrobial activity and protease stability. The antimicrobial mechanism studied revealed that the MDLs showed a rapid bactericidal effect by interacting with lipopolysaccharide (LPS) or lipoteichoic acid (LTA) and penetrating the bacterial cell membrane. In conclusion, we designed and synthesized a new class of MDLs with potent antimicrobial activity, high proteolytic stability, and low hemolytic activity through N-terminal fatty acid conjugation. Full article

The production of therapeutic recombinant toxins requires careful host cell selection. Bacteria, yeast, and mammalian cells are common choices, but no universal solution exists. Achieving the delicate balance in toxin production is crucial due to potential self-intoxication. Recombinant toxins from various sources find applications in antimicrobials, biotechnology, cancer drugs, and vaccines. “Toxin-based therapy” targets diseased cells using three strategies. Targeted cancer therapy, like antibody–toxin conjugates, fusion toxins, or “suicide gene therapy”, can selectively eliminate cancer cells, leaving healthy cells unharmed. Notable toxins from various biological sources may be used as full-length toxins, as plant (saporin) or animal (melittin) toxins, or as isolated domains that are typical of bacterial toxins, including Pseudomonas Exotoxin A (PE) and diphtheria toxin (DT). This paper outlines toxin expression methods and system advantages and disadvantages, emphasizing host cell selection’s critical role. Full article

With the increasing challenge of controlling infectious diseases due to the emergence of antibiotic-resistant strains, the importance of discovering new antimicrobial agents is rapidly increasing. Animal venoms contain a variety of functional peptides, making them a promising platform for pharmaceutical development. In this study, a novel toxin peptide with antibacterial and anti-inflammatory activities was discovered from the spider venom gland transcriptome by implementing computational approaches. Lycotoxin-Pa2a (Lytx-Pa2a) showed homology to known-spider toxin, where functional prediction indicated the potential of both antibacterial and anti-inflammatory peptides without hemolytic activity. The colony-forming assay and minimum inhibitory concentration test showed that Lytx-Pa2a exhibited comparable or stronger antibacterial activity against pathogenic strains than melittin. Following mechanistic studies revealed that Lytx-Pa2a disrupts both cytoplasmic and outer membranes of bacteria while simultaneously inducing the accumulation of reactive oxygen species. The peptide exerted no significant toxicity when treated to human primary cells, murine macrophages, and bovine red blood cells. Moreover, Lytx-Pa2a alleviated lipopolysaccharide-induced inflammation in mouse macrophages by suppressing the expression of inflammatory mediators. These findings not only suggested that Lytx-Pa2a with dual activity can be utilized as a new antimicrobial agent for infectious diseases but also demonstrated the implementation of in silico methods for discovering a novel functional peptide, which may enhance the future utilization of biological resources. Full article

Methicillin-resistant Staphylococcus aureus (MRSA) often cause infections with high mortality rates. Antimicrobial peptides are a source of molecules for developing antimicrobials; one such peptide is melittin, a fraction from the venom of the Apis mellifera bee. This study aimed to evaluate the antibacterial and antibiofilm activities of melittin and its association with oxacillin (mel+oxa) against MRSA isolates, and to investigate the mechanisms of action of the treatments on MRSA. Minimum inhibitory concentrations (MICs) were determined, and synergistic effects of melittin with oxacillin and cephalothin were assessed. Antibiofilm and cytotoxic activities, as well as their impact on the cell membrane, were evaluated for melittin, oxacillin, and mel+oxa. Proteomics evaluated the effects of the treatments on MRSA. Melittin mean MICs for MRSA was 4.7 μg/mL and 12 μg/mL for oxacillin. Mel+oxa exhibited synergistic effects, reducing biofilm formation, and causing leakage of proteins, nucleic acids, potassium, and phosphate ions, indicating action on cell membrane. Melittin and mel+oxa, at MIC values, did not induce hemolysis and apoptosis in HaCaT cells. The treatments resulted in differential expression of proteins associated with protein synthesis and energy metabolism. Mel+oxa demonstrated antibacterial activity against MRSA, suggesting a potential as a candidate for the development of new antibacterial agents against MRSA. Full article

Viruses are known to infect most types of organisms. In humans, they can cause several diseases that range from mild to severe. Although many antiviral therapies have been developed, viral infections continue to be a leading cause of morbidity and mortality worldwide. Therefore, the discovery of new and effective antiviral agents is desperately needed. Animal venoms are a rich source of bioactive molecules found in natural goods that have been used since ancient times in alternative medicine to treat a variety of human diseases. Recently, and with the onset of the COVID-19 pandemic, scientists have regained their interest in the possible use of natural products, such as bee venom (BV), as a potential antiviral agent to treat viral infections. BV is known to exert many therapeutic activities such as anti-proliferative, anti-bacterial, and anti-inflammatory effects. However, there is limited discussion of the antiviral activity of BV in the literature. Therefore, this review aims to highlight the antiviral properties of BV and its two primary constituents, melittin (MEL) and phospholipase A2 (PLA2), against a variety of enveloped and non-enveloped viruses. Finally, the innovative strategies used to reduce the toxicity of BV and its two compounds for the development of new antiviral treatments are also considered. Full article

Antimicrobial peptides are key components of the immune system. These peptides affect the membrane in various ways; some form nano-sized pores, while others only produce minor defects. Since these peptides are increasingly important in developing antimicrobial drugs, understanding the mechanism of their interactions with lipid bilayers is critical. Here, using atomic force microscopy (AFM), we investigated the effect of a synthetic hybrid peptide, CM15, on the membrane surface comprising E. coli polar lipid extract. Direct imaging of supported lipid bilayers exposed to various concentrations of the peptide revealed significant membrane remodeling. We found that CM15 interacts with supported lipid bilayers and forms membrane-spanning defects very quickly. It is found that CM15 is capable of remodeling both leaflets of the bilayer. For lower CM15 concentrations, punctate void-like defects were observed, some of which re-sealed themselves as a function of time. However, for CM15 concentrations higher than 5 µM, the defects on the bilayers became so widespread that they disrupted the membrane integrity completely. This work enhances the understanding of CM15 interactions with the bacterial lipid bilayer. Full article