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The previously described decapeptide AKVTMTCSAS (killer peptide, KP), derived from the variable region of a recombinant yeast killer toxin-like anti-idiotypic antibody, proved to exert a variety of antimicrobial, antiviral, and immunomodulatory activities. It also showed a peculiar self-assembly ability, likely responsible for the therapeutic effect in animal models of systemic and mucosal candidiasis. The present study analyzed the biological and structural properties of peptides derived from KP by substitution or deletion of the first residue, leaving unchanged the remaining amino acids. The investigated peptides proved to exert differential in vitro and/or in vivo anti-Candida activity without showing toxic effects on mammalian cells. The change of the first residue in KP amino acidic sequence affected the conformation of the resulting peptides in solution, as assessed by circular dichroism spectroscopy. KP-derivatives, except one, were able to induce apoptosis in yeast cells, like KP itself. ROS production and changes in mitochondrial transmembrane potential were also observed. Confocal and transmission electron microscopy studies allowed to establish that selected peptides could penetrate within C. albicans cells and cause gross morphological alterations. Overall, the physical and chemical properties of the first residue were found to be important for peptide conformation, candidacidal activity and possible mechanism of action. Small antimicrobial peptides could be exploited for the development of a new generation of antifungal drugs, given their relative low cost and ease of production as well as the possibility of devising novel delivery systems. [ABSTRACT FROM AUTHOR]

Background: A peptide of 2,733 Da named SP-E, previously isolated from pig saliva and already described for its antifungal activity and absence of toxicity against mammalian cells, is characterized by a high content of proline residues (70% of entire sequence), that confer structural features probably related to peptide activity. Purpose: The aim of this study was to evaluate the activity of SP-E against Gram-negative bacteria, including drug-resistant clinical isolates. Methods: SP-E and shorter fragments of the same peptide were tested in vitro against the selected bacteria by colony forming unit assays. Scanning electron microscopy and confocal microscopy were also applied. SP-E potential therapeutic activity was evaluated in vivo in a Galleria mellonella model of bacterial infection. Results: SP-E proved to be active against the tested bacteria with EC50 values in the micromolar range. Though maintaining antibacterial properties, the shorter peptides showed lower activity in respect to the parental molecule. Kinetics of killing action and nonmembranolytic internalization within Escherichia coli and Pseudomonas aeruginosa cells strongly suggested a cytosolic mechanism of action involving one or more intracellular molecular targets. A single injection of SP-E exerted a therapeutic effect in G. mellonella larvae infected with P. aeruginosa. Conclusion: The biological properties of SP-E strongly back this peptide as a new promising multitasking antimicrobial molecule. [ABSTRACT FROM AUTHOR]

Enterococcus faecalis is a common cause of biofilm-associated opportunistic infections, which are often difficult to treat. The formation of E. faecalis biofilms on the dentinal walls of the root canal is frequently the cause of endodontic treatment failure and secondary apical periodontitis. In a preliminary work, two recognized antifungal peptides, KP and L18R, showed antibacterial activity against planktonic E. faecalis cells at micromolar concentrations. Moreover, L18R proved to reduce the biomass in the early stage of E. faecalis biofilm development on polystyrene plates, while a qualitative biofilm inhibition was demonstrated on hydroxyapatite disks by confocal laser scanning microscopy (CLSM). The aim of this study was to better characterize the effect of both peptides on E. faecalis biofilm. A reduction in metabolic activity after peptide treatment was detected by Alamar Blue assay, while a remarkable impairment in the architecture of E. faecalis biofilms on hydroxyapatite disks, along with a significant reduction in viable bacteria, was caused mostly by L18R, as assessed by CLSM and scanning electron microscopy. The lack of cytotoxicity of the investigated peptides against L929 murine fibroblasts was also determined. Obtained results suggest L18R as a promising candidate for the development of new strategies for endodontic infection control. [ABSTRACT FROM AUTHOR]

The growing problem of antimicrobial resistance highlights the need for alternative strategies to combat infections. From this perspective, there is a considerable interest in natural molecules obtained from different sources, which are shown to be active against microorganisms, either alone or in association with conventional drugs. In this paper, peptides with the same sequence of fragments, found in human serum, derived from physiological proteins, were evaluated for their antifungal activity. A 13-residue peptide, representing the 597–609 fragment within the albumin C-terminus, was proved to exert a fungicidal activity in vitro against pathogenic yeasts and a therapeutic effect in vivo in the experimental model of candidal infection in Galleria mellonella. Studies by confocal microscopy and transmission and scanning electron microscopy demonstrated that the peptide penetrates and accumulates in Candida albicans cells, causing gross morphological alterations in cellular structure. These findings add albumin to the group of proteins, which already includes hemoglobin and antibodies, that could give rise to cryptic antimicrobial fragments, and could suggest their role in anti-infective homeostasis. The study of bioactive fragments from serum proteins could open interesting perspectives for the development of new antimicrobial molecules derived by natural sources. [ABSTRACT FROM AUTHOR]

The synthetic peptide T11F (TCRVDHRGLTF), derived from the constant region of human IgM antibodies, proved to exert a significant activity in vitro against yeast strains, including multidrug resistant isolates. Alanine substitution of positively charged residues led to a decrease in candidacidal activity. A more dramatic reduction in activity resulted from cysteine replacement. Here, we investigated the conformational properties of T11F and its alanine-substituted derivatives by circular dichroism (CD) and nuclear magnetic resonance (NMR) spectroscopy. Peptide interaction with Candida albicans cells was studied by confocal and scanning electron microscopy. T11F and most of its derivatives exhibited CD spectra with a negative band around 200 nm and a weaker positive band around 218 nm suggesting, together with NMR coupling constants, the presence of a polyproline II (PPII) helix, a conformational motif involved in a number of biological functions. Analysis of CD spectra revealed a critical role for phenylalanine in preserving the PPII helix. In fact, only the F11A derivative presented a random coil conformation. Interestingly, the loss of secondary structure influenced the rate of killing, which turned out to be significantly reduced. Overall, the obtained results suggest that the PPII conformation contributes in characterising the cell penetrating and fungicidal properties of the investigated peptides. [ABSTRACT FROM AUTHOR]

Abstract: A proline-rich peptide of 2733Da, isolated from pig parotid granule preparations was tested against different pathogenic fungi. It showed interesting antifungal activity towards a clinical isolate of Cryptococcus neoformans, with an EC50 of 2.2μM. Neither cytotoxic nor haemolytic effects were observed towards mammalian cells. Circular dichroism and infrared spectroscopic studies showed that the peptide adopted a combination of polyproline type-II, β-turn and unordered conformations at physiological temperatures. Temperature dependent experiments evidenced a tendency to adopt a polyproline-II helix conformation. From experiments with lipid vesicles, Neutral Red Uptake (NRU), haemolytic assays, and confocal microscopy studies, it could be hypothesized that the peptide may exert its antifungal effect by interacting with an intracellular target rather than through membrane damage. [Copyright &y& Elsevier]