Letícia Oliveira Catarin Nunes

Background/Objectives: Tuberculosis remains a major public health challenge due to the persistence of Mycobacterium tuberculosis (Mtb) and the emergence of multidrug-resistant strains. In this study, Pep-H, an HNP-1-derived antimicrobial peptide with the sequence RRYGTCIYQGRLWAF-NH₂, was used as a compact scaffold to examine how single-residue substitutions at the Cys position affected its biological and functional profile. Methods: A focused single-position substitution panel was generated by replacing Cys with Trp, Ala, Arg, or Met while preserving peptide length and sequence context, and the analogs were computationally prioritized according to their predicted antitubercular potential and contrasting side-chain properties. The peptides were synthesized, purified, characterized by HPLC and mass spectrometry, and evaluated for activity against Mtb H37Rv, cytotoxicity, hemolysis, ethidium bromide accumulation, and DPPH radical scavenging. Results: Pep-H retained the most favorable profile, showing the highest antimycobacterial potency, low hemolysis, favorable selectivity indices, enhanced ethidium bromide accumulation, and the strongest antioxidant response. All Cys substitutions reduced antimycobacterial activity, indicating that none of the tested residues reproduced the integrated biological profile of Pep-H. Conclusions: The contrasting outcomes of the Arg- and Met-containing analogs suggest that increased cationicity or sulfur retention alone was insufficient, while supporting a multifactorial contribution of Cys side-chain chemistry and the local GTCIY environment.

Antimicrobial peptides such as Melittin exhibit potent broad-spectrum activity but are limited by high cytotoxicity. The rational design of bioinspired Melittin-derived analogues represents a promising strategy to reduce toxicity while maintaining antimicrobial efficacy. In this study, Melittin and analogues (TT-1, FKW, and WKW) were synthesized using solid-phase peptide synthesis (SPPS) and characterized for biological and biophysical essays. Antimicrobial and hemolytic activity, serum stability, secondary structure, and membrane interaction were analysed. FKW and WKW exhibited broad-spectrum antimicrobial activity, with minimum inhibitory concentration (MIC) as low as 8 and 16 µg/mL against Staphylococcus aureus and Enterococcus faecium. Both analogues also showed improved activity against Klebsiella pneumoniae (32 µg/mL) and Pseudomonas aeruginosa (128 and 256 µg/mL for FKW and WKW, respectively) compared to Melittin (64 and 512 µg/mL). In terms of cytotoxicity, FKW and WKW showed significantly reduced hemolytic activity, with HC₅₀ values of 264.8 µg/mL and 237.2 µg/mL, respectively, resulting in improved selectivity indexes relative to Melittin (HC₅₀ of 7.9 µg/mL). In liposomes, both adopt α-helical structures and cause disruption via pore formation or detergent-like mechanisms. TT-1 showed minimal toxicity but weak antimicrobial activity (MIC > 256 µg/mL). Although FKW and WKW exhibited limited serum stability (half-lives of 2.2 and 1.5 h), their degradation may reduce systemic toxicity. Overall, these analogues demonstrate an improved balance between antimicrobial activity and safety.

Tuberculosis (TB), caused by Mycobacterium tuberculosis ( Mtb ), remains a major global health concern, particularly due to the emergence of multidrug‐resistant and extensively drug‐resistant strains. The persistence and propagation of TB are favored by the pathogen's sophisticated virulence mechanisms, its ability to evade immune responses, and the formation of latent infections within granulomas. Current therapeutic regimens are limited by long treatment durations, drug resistance, and significant socioeconomic burdens. Antimicrobial peptides (AMPs) have emerged as promising alternatives because of their broad‐spectrum activity and reduced likelihood of resistance development. Nevertheless, their clinical application is hindered by rapid proteolytic degradation, low specificity and limited bioavailability. Recent advances in nanotechnology have facilitated the encapsulation and targeted delivery of AMPs, improving their therapeutic potential against TB. Furthermore, the integration of computational approaches—such as molecular docking and molecular dynamics (MD) simulations—has enabled the rational design and optimization of AMPs, expediting the discovery of novel anti‐TB agents. This review summarizes the pathogenesis and resistance mechanisms of Mtb , highlights the current landscape and limitations of AMP‐based therapies, and discusses the role of nanotechnology and in silico tools in the development of new treatment strategies for TB.

Although peptides have been shown to have biological functions in neurodegenerative diseases, their role in Parkinson’s disease has been understudied. A previous study by our group, which used a 6-hydroxydopamine zebrafish model, suggested that nine intracellular peptides may play a part in this condition. In this context, our aim is to better understand the role of five of these nine peptides. The selection of peptides was made based on their precursor proteins, which are fatty acid binding protein 7, mitochondrial ribosomal protein S36, MARCKS-related protein 1-B, excitatory amino acid transporter 2 and thymosin beta-4. The peptides were chemically synthesized in solid phase and characterized by high-performance liquid chromatography and mass spectrometry. Circular dichroism was performed to determine the secondary structure of each peptide, which showed that all five peptides maintain a random structure in the aqueous solutions that were studied. Two molecules show a helical profile in trifluoroethanol, a known structuring agent. Cell viability by the MTT assay indicates that all five peptides are not cytotoxic in all concentrations tested in both mouse and human cell lines. Behavioral assay using a 6-OHDA zebrafish larvae model suggest that all peptides help in the recovery of motor function with 24 h treatment at two concentrations. Three peptides showed a complete recovery from the 6-OHDA-induced motor impairment. Further studies are needed to better understand the mechanism of action of these peptides and whether they are truly a potential ally against Parkinson’s disease.

Currently, the search for new alternatives to conventional antibiotics to combat bacterial resistance is an urgent task, as many microorganisms threaten human health due to increasing bacterial resistance to traditional medicines. Thus, new molecules such as antimicrobial peptides have emerged as promising alternatives because of their low induction of resistance and broad spectrum of action. In this context, in the past few years, our research group has synthesized and characterized a peptide derived from the C-terminal region of the Lys⁴⁹ PLA₂-like BthTX-I, named p-BthTX-I. After several studies, the peptide (p-BthTX-I)₂K was proposed as the molecule with the most considerable biotechnological potential. As such, the present work aimed to evaluate whether the modifications made on the peptide (p-BthTX-I)₂K can be applied to other molecules originating from the C-terminal region of PLA₂-like Lys⁴⁹ from snake venoms. The peptides were obtained through the solid-phase peptide synthesis technique, and biochemical and functional characterization was carried out using dichroism techniques, mass spectrometry, antimicrobial activity against ESKAPE strains, hemolytic activity, and permeabilization of lipid vesicles. The antimicrobial activity of the peptides was promising, especially for the peptides (p-AppK)₂K and (p-ACL)₂K, which demonstrated activity against all strains that were tested, surpassing the model molecule (p-BthTX-I)₂K in most cases and maintaining low hemolytic activity. The modifications initially proposed for the (p-BthTX-I)₂K peptide were shown to apply to other peptides derived from Lys⁴⁹ PLA₂-like from snake venoms, showing promising results for antimicrobial activity. Future assays comparing the activity of the dimers obtained through this strategy with the monomers of these peptides should be carried out.