Cancer is a major cause of death worldwide, and despite advancements in treatment, no effective cure exists. Pro-apoptotic peptides have emerged as a promising alternative to conventional anticancer drugs. However, their clinical application faces challenges due to the cell membrane's hydrophobic barrier, limiting peptide access to intracellular targets. To overcome this obstacle, research suggests that conjugating with cell-penetrating peptides (CPPs) can improve intracellular transport. We propose a bioinformatics approach to evaluate the efficiency of four CPPs (TAT, R8, ATP 128, and Penetramax) for delivering pro-apoptotic sequences BIM, NOXA, BID, and BMF. We also used four anti-cancer peptides (LL37, Pexig, CLS001, and Magainin II) to enhance CPP potency. Our method resulted in the selection of 60 CppProAcp sequences with a high and average probability of absorption efficiency. We evaluated these peptides (CppProAcp) for transfer effect, stability, thermodynamic properties, aggregation potential, folding speed, backbone flexibility, and in vivo administration sensitivity, resulting in the identification of 20 promising structures. Based on our study, this bioinformatic workflow can be universally applied to any CPP-peptide conjugation scheme.

The innate immune system of most vertebrates includes a family of host defense peptides named cathelicidins. Avian cathelicidins are excellent candidates for antimicrobial drug development due to their broad spectrum of activity against various microorganisms and low toxicity. Because of their extraordinarily long lifespan, parrots are exposed to a variety of infectious diseases during their lifetime, making them an excellent subject for the study of their immune system. The aim of this study was to evaluate the antimicrobial, hemolytic and cytotoxic activity of parrot cathelicidins and to determine their mode of action. To conduct in vitro tests, we synthesized six peptides: two from Amazona guildingii, two from Eolophus roseicapilla, and two from Gallus gallus (as reference peptides). Our data demonstrated that even at the greatest concentration examined, the peptides had no effect on VERO cells and human erythrocytes. At lower concentrations, the peptides showed strong antimicrobial activity with MIC and MBC values ranging from 12.5 µM to 1.56 µM. However, none of the peptides showed activity against Candida albicans. We used Sytox Green and performed a 4-hours time-kill assay to investigate the mechanism of action. The kinetic assay showed that the peptides required between 20 and 60 min to kill Escherichia coli and that they operated by disturbing the bacterial membrane. In general, parrot cathelicidin-derived peptides showed potent antimicrobial activity and could be used as novel templates for antimicrobial drug development.

Cathelicidins are a family of host defense peptides (HDP) with different functions, highlighting their antimicrobial activity, expressed in most vertebrates. These peptides present a high variability, differentiating their sequence, structure, and function among different animal species. In this research, mining the available reptile genomes on NCBI database has been carried out to identify new cathelicidins. An analysis of the structure and physicochemical properties of the peptides was made and 6 of them were selected to synthesize and study their potential as antimicrobial compounds in vitro. For this, the minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC) were characterized against 5 human pathogenic bacteria. For peptides that showed the highest activity, their bacteriostatic or bactericidal character as well as their lytic activity and their period of action was studied. For this, kill curves were carried out with the bacteria in the latency and exponential phase. To study the possible toxicity of peptides, a hemolytic activity assay with rat erythrocytes and a MTT cytotoxicity assay with Vero cells were carried out. Novel reptile peptides showed good antimicrobial activity between 0.39 and 25 micromolar against the bacteria which they were tested, exhibiting bactericidal activity when the microorganism is in the latency phase and bacteriostatic activity when it's in the exponential phase. Peptides had an active period of approximately 6 hours and showed lytic activity against bacteria. On the other hand, there was no hemolytic or cytotoxic activity at concentrations that they were effective against microorganisms.

Fish constantly interact with microorganisms in their aquatic habitat. As a defense mechanism, they possess innate immune system-derived antimicrobial peptides that combat bacteria, viruses, and fungi. The piscidin family's characterization has been limited to Teleostei fish. This study aimed to select a subset of peptides annotated as piscidins in NCBI and evaluate their bioactivity and structure through both in silico and in vitro methodologies.

Beginning with 51 piscidin sequences, bioinformatic tools were utilized to screen for potential active peptides. These peptides exhibited attributes like α-helical structure, cationic charge, and potent activity against microorganisms and tumors. To explore promising candidates, five novel piscidin peptides were synthesized alongside a well-established active peptide, Epinecidin-1 from Epinephelus coioides. Our initial focus was assessing antibiotic and antifungal activities of each peptide against human pathogens (Escherichia coli, Staphylococcus aureus, Salmonella enterica, Enterococcus faecalis, Pseudomonas aeruginosa, Campylobacter jejuni, and Candida albicans). Outcomes demonstrated noteworthy activity at low concentrations, e.g., 1.56 µM against S. aureus or 3.125-6.25 µM against P. aeruginosa. Peptide toxicity was evaluated using rat erythrocytes and the Vero cell line, revealing toxicity at higher concentrations (25-50 µM) with milder effects at lower concentrations.

In conclusion, these peptides exhibited substantial in vitro antimicrobial activity, particularly against bacteria, aligning with in silico forecasts. These findings strongly underscore the potential of these peptides as agents against microorganisms, potentially aiding in the fight against antibiotic resistance. However, further research is imperative to delve into their mode of action and other potential immunomodulatory attributes.

Antimicrobial peptides (AMPs), a class of innate immune molecules, have received significant attention for their capacity to combat a broad spectrum of pathogens, including viruses, bacteria, and fungi. Predicting AMPs has made it easy and efficient to find AMPs from large datasets with high accuracy. Recent years have witnessed wide applications of computational methods especially machine learning and deep learning for discovering and engineering AMPs. However, existing methods only use features including compositional, physiochemical, and structural properties of peptide sequences, which cannot fully capture information from AMPs. Here, we present SAMP, an ensemble random projection (RP) based computational model that leverages a new type of features called proportionalized split amino acid composition (PSAAC) in addition to conventional sequence-based features for AMP prediction. With this new feature set, SAMP captures the residue patterns like sorting signals at around both the N terminus and C terminus, while also retaining the sequence order information from the middle peptide fragments. Benchmarking tests on balanced and imbalanced datasets from different species demonstrate that SAMP consistently outperforms existing state-of-the-art methods, such as iAMPpred, in terms of accuracy, sensitivity, specificity and AUC. We further incorporate the ensemble RP architecture in our model, so that our model SAMP is scalable to processing large scale AMP screening with further performance improvement, compared to those without RP. To enhance the impact of SAMP, we have developed a Python package for it, which is freely and publicly available at https://github.com/wan-mlab/SAMP.

The antibacterial effects of antimicrobial peptides (AMPs) are related to their ability to disrupt bacterial membranes. Understanding the interaction between AMPs and membranes helps the active peptide design. Molecular dynamics simulations are a powerful tool to access the atomic-level scale of the peptide interaction with membranes. Somuncurin-1 is a small AMP isolated from the Patagonian frog Pleurodema somuncurense that showed moderate antimicrobial activity against E. coli and S. aureus, along with low cytotoxicity. To propose modifications in the somuncurin-1 sequence that could increase its performance, we studied its behavior against two membrane models using molecular dynamics simulations. Two types of lipid bilayers were considered: mixtures of palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and lipid mix of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoglycerol (POPG) and 1-palmitoyl-2-oleoylphosphatidylethanolamine (POPE), which are simple models to mimic mammalian and bacterial membranes, respectively. The results show a differential behavior on these membranes. We found cooperative effects in the peptide-lipid bilayer interaction. Somuncurin-1 was distributed at the hydrophobic core-water interface, and simulations were conducted at temperatures of 303 K, 310 K, and 320 K. Somuncurin-1 exhibited distinct behavior in bilayer-peptide interaction: they display a strong affinity for the lipid interface of the bacterial model. Lys residue is found to anchor at the lipid water interface due to electrostatic interactions with the lipid heads. These simulations provide valuable insights into the behavior somuncurin-1 as a function of its concentration, temperature, and membrane composition in its interactions with lipid bilayers, offering a foundation for optimizing its therapeutic potential.

The search for innovative therapeutic solutions to address the escalating resistance of pathogens to antibiotics is a pressing global challenge. Bioprospecting for novel molecules is a cornerstone, providing essential data to fuel databases used in developing AI-driven tools. These tools are crucial for drug design, ushering in a new drug discovery and development era. Antimicrobial peptides (AMPs) have demonstrated their effectiveness against a broad spectrum of pathogens. These peptides are sourced from various natural origins, including amphibian skin, which is particularly abundant. The northern region of Argentina boasts an unexplored and diverse amphibian biodiversity, promising potential for the discovery of new AMPs. This study aimed to identify AMPs from Leptodactylus chaquensis frog skin collected in Corrientes, Argentina. Total mRNA was extracted from the skin, cDNA was synthesized, and cloned into E. coli DH5α cells. Plasmids of selected colonies were purified, PCR amplified, insert size assessed, and sequences obtained. Three prepro-peptides coding novel mature peptides were identified. The peptides, provisionally named Lch-1, Lch-2, and Lch-3, exhibited lengths of 25, 21, and 9 residues, with net charges of +1, 0, and -1, respectively. Hydrophobic percentages were 38%, 54%, and 44%, with theoretical 3D α-helical content in their structures. Lch-1 and -2 shared 43% and 41% sequence similarity with ocellatin-4 and -6 from Leptodactyus ocellatus. Predicted interactions with membranes suggest antimicrobial potential using the APD tool. Peptides will be synthesized for bioactivity assays. These findings underscore the significance of bioprospecting in uncovering distinct and unique bioactive compounds within each species.

Antimicrobial peptides (AMPs) are small proteins that play an important role in the innate immune system of various organisms, including plants, animals, and humans. These natural defence molecules have attracted considerable interest due to their potential as alternative antimicrobial agents to combat infectious diseases. In this study, we used computational and in vitro methods to investigate the antimicrobial activity of cathelicidin family peptides from bat species with different ecological niches.

The study of the physicochemical parameters of the peptides (hydrophobicity and net charge), together with the study of the helical regions, allowed us to deduce the antimicrobial character of peptides. To analyse the antimicrobial activity in vitro, we first determine the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) against four different bacteria: E. coli, Salmonella, S. aureus, and E. faecalis. We then selected the most effective peptide and assessed whether it acts as a bacteriostatic or bactericidal agent. Additionally, we investigated the duration of its activity and its ability to lyse bacterial cells. We accomplished this by plotting killing curves during both the exponential and latent phases of bacterial growth. Finally, we evaluated the peptide's potential to cause hemolysis in rat erythrocytes. 10⁵

One of our peptides revealed a high antimicrobial activity, with MIC and MBC values ranging from 3.12 to 1.56 μM. It also demonstrated bactericidal properties during the stationary phase but acted as a bacteriostatic agent during the exponential phase. Notably, it exhibited lytic activity against the tested bacteria but no hemolytic activity against rat erythrocytes.

Proline-rich antimicrobial peptides (PrAMPs) are short peptides naturally synthesized by arthropods and mammals. According to their ability to inhibit translation, PrAMPs are classified into two subgroups. While type I PrAMPs arrest ribosomes at start codon of open reading frames, type II PrAMPs apidaecin (Api) and drosocin (Dro) stall ribosomes at stop codons. Notably, Api and Dro share sequence similarities specially at their C-termini, which include amino acids critical for target interaction. In this work, we bioinformatically identified Dro-like peptides encoded in the genomes of diverse fruit fly species. By testing the antimicrobial activity of the synthetic version of ten of these peptides, we found that only two of them (besides the already characterized Dro from D. melanogaster) were able to kill the tested Gram-negative bacteria. Further, the identified active peptides had the ability to cause stop codon readthrough in E. coli cells and cause ribosome arrest at stop codons during in vitro translation. Intriguingly, the peptides unable to penetrate and kill the tested strains arrested ribosomes at start codons, resembling the behavior of type I PrAMPs. The outcomes of our study underscore the complexity of predicting MOA solely based on similarity of peptide sequences.

We have shown that LL-37, its truncated (GI-20, GF-17) and engineered (17BIPHE2) forms possess spermicidal effects and microbicidal activity against Neisseria gonorrhoeae. However, 17BIPHE2 has the highest spermicidal activity on human sperm resuspended in cervicovaginal fluid-containing medium due to its resistance to proteases. 17BIPHE2 should therefore be developed into a vaginal MPT agent. However, <5% of 17BIPHE2/LL-37/GI-20/GF-17 remained in the mouse reproductive tract after its intravaginal administration. A proper delivery formulation is thus needed. We have chosen the universal placebo gel, hydroxyethylcellulose (HEC), as an excipient in 17BIPHE2 formulation. HEC has been used safely as a vaginal lubricant and the uterus of mice injected with 2% HEC was still normal. 17BIPHE2 (32.4 or 86.4 µM) was solubilized in 2% HEC in a bicarbonate-5% CO₂ buffered isotonic salt solution (HEC-BS). 17BIPHE2-HEC-BS possessed in vitro spermicidal activity on mouse and human sperm. The contraceptive effects of 17BIPHE2-HEC-BS were then evaluated in mice. Since semen ejaculated in the mouse vagina is immediately swept into the uterine lumen, 17BIPHE2-HEC-BS was transcervically administered. Immunoblotting of uterine fluid collected from mice sacrificed 3 hours after transcervical injection indicated that 17BIPHE2 was present in a substantial amount, but the amount was decreased ~50% 24 hours after the administration. Through reaction with orcinol, HEC was shown to be present in the uterine fluid with amounts temporally declined like 17BIPHE2. Pregnancy of mice administered with 17BIPHE2 (86.4 µM)-HEC-BS was then determined. Only three of seven mice (43%) cycling in estrus, which were transcervically injected with 17BIPHE2-HEC-BS and then co-caged with fertile males for 3 days, became pregnant. In contrast, 100% pregnancy was observed in mice transcervically injected with HEC alone (n=5). While the results were promising, the experiments need to be done with more mice with modifications (e.g., with one-day co-caging with males and/or different percentages of HEC in formulation).