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Candida albicans is an opportunistic fungal pathogen and a commensal organism that commonly colonizes mucosal surfaces, including those inside the human mouth. To help control C. albicans, human saliva contains the antifungal peptide histatin 5 (Hst‐5), which has strong antifungal activity against C. albicans. However, the pathogen produces secreted aspartic proteases (Saps) that cleave Hst‐5 at lysine residues and eliminate its antifungal properties. We designed variants of Hst‐5 with its lysine residues substituted with arginine or leucine to evaluate the effect on proteolysis by Saps. We found site‐, residue‐, and Sap‐dependent effects from single amino acid substitutions. The K17R and K17L modifications led to dramatic results, with over 77% and 100% intact peptide remaining after incubation with Sap9 and Sap2, respectively, compared to 47% and 61% of Hst‐5. This decrease in proteolysis was accompanied by a reduction in cleavage on the C‐terminal side of K17, suggesting the Saps prefer lysine at K17 for cleavage. Incubation with C. albicans cells and culture supernatant corroborated the results with purified Saps and highlighted their biological relevance. The modifications to Hst‐5 do not diminish the antifungal activity of Hst‐5, and, in fact, the K17R, K17L, and K11R peptides retained significantly more antifungal activity after treatment with Saps than Hst‐5. Our results indicate that single amino acid modifications drastically impact both proteolysis at the modification sites and the overall level of proteolysis of the peptide, demonstrating the potential of designing peptides for resistance to proteolysis as a means for improving therapeutic efficacy. [ABSTRACT FROM AUTHOR]

Histatin5 (Hst-5) is a member of the histatin family of antimicrobial peptides secreted by human parotid and submandibular glands. With the natural antibacterial activity, it plays an important role in the first-line barrier of oral cave against pathogens, especially for the fungal intrusion. In this study, we explored the utility of Hst-5 in the treatment of vulvovaginal candidiasis, a common condition of women of the childbearing age. We used a synthesized Hst-5 over five consecutive days as the topical treatment in a murine model of vulvovaginal candidiasis. According to the fungal colony counts, fungal burden in the vagina lavage dropped remarkably after treatment with Hst-5. Furthermore, cytological analysis of the lavage fluid indicated that the number of cast-off cells including cornified epithelial cells and inflammatory cells also decreased; histological evaluation of the vagina tissue revealed less fungi adhering to the vaginal wall in treated animals than in controls. Combined, these results suggested for the first time the potential utility of Hst-5 as a topical treatment for vulvovaginal candidiasis, uncovering the possibility of exploiting the natural antibiotic peptides in other aspects. [ABSTRACT FROM AUTHOR]

Background: Histatins are histidine rich polypeptides produced in the parotid and submandibular gland and secreted into the saliva. Histatin-3 and -5 are the most important polycationic histatins. They possess antimicrobial activity against fungi such as Candida albicans. Histatin-5 has a higher antifungal activity than histatin-3 while histatin-3 is mostly involved in wound healing in the oral cavity. We found that these histatins, like other polycationic peptides and proteins, such as LL-37, lysozyme and histones, interact with extracellular actin. Results: Histatin-3 and -5 polymerize globular actin (G-actin) to filamentous actin (F-actin) and bundle F-actin filaments. Both actin polymerization and bundling by histatins is pH sensitive due to the high histidine content of histatins. In spite of the equal number of net positive charges and histidine residues in histatin-3 and -5, less histatin-3 is needed than histatin-5 for polymerization and bundling of actin. The efficiency of actin polymerization and bundling by histatins greatly increases with decreasing pH. Histatin-3 and -5 induced actin bundles are dissociated by 100 and 50 mM NaCl, respectively. The relatively low NaCl concentration required to dissociate histatin-induced bundles implies that the actin-histatin filaments bind to each other mainly by electrostatic forces. The binding of histatin-3 to F-actin is stronger than that of histatin-5 showing that hydrophobic forces have also some role in histatin-3-actin interaction. Histatins affect the fluorescence of probes attached to the D-loop of Gactin indicating histatin induced changes in actin structure. Transglutaminase cross-links histatins to actin. Competition and limited proteolysis experiments indicate that the main histatin cross-linking site on actin is glutamine-49 on the D-loop of actin. Conclusions: Both histatin-3 and -5 interacts with actin, however, histatin 3 binds stronger to actin and affects actin structure at lower concentration than histatin-5 due to the extra 8 amino acid sequence at the C-terminus of histatin-3. Extracellular actin might regulate histatin activity in the oral cavity, which should be the subject of further investigation. [ABSTRACT FROM AUTHOR]

INTRODUCTION: The mechanisms of adhesion to solid surfaces enable S. mutans to colonize oral cavities and form biofilms, which play an important role in caries development. Additional properties enabling the survival of S. mutans in the oral cavity include its ability to survive in acidic environments and specific interactions with other microorganisms inhabiting this ecosystem. AIM OF THE STUDY: The aim of this study was to determine the antibacterial activity of saliva histatin-5 (peptide) and lysozyme (protein) against S. mutans and L. rhamnosus, as representatives of physiological flora. MATERIALS AND METHODS: The study involved strains of physiological (L. rhamnosus) and cariogenic (S. mutans) flora isolated from one patient with diagnosed early caries of the deciduous teeth. RESULTS: It was proved that the presence of probiotic L. rhamnosus bacteria in the environment had a negative impact on the ability of S. mutans to produce biofilm. Moreover, the antibacterial activity of histatin-5 was confirmed, and it inhibited S. mutans growth at concentrations of 27.2 μg/ml and 54.4 μg/ml, both individually and in a mixture with lysozyme (in a total concentration of 54.4 μg/ml). CONCLUSIONS: The data obtained constitute a promising result due to their potential future application in the prevention and early diagnosis of caries. [ABSTRACT FROM AUTHOR]

Histatin-5 (Hst-5, DSHAKRHHGYKRKFHEKHHSHRGY) is a member of a histidine-rich peptide family secreted by major salivary glands, exhibiting high fungicidal activity against Candida albicans. In the present work, we demonstrate the 3D structure of the head-to-tail cyclic variant of Hst-5 in TFE solution determined using NMR spectroscopy and molecular dynamics simulations. The cyclic histatin-5 reveals a helix-loop-helix motif with α-helices at positions Ala4-His7 and Lys11-Ser20. Both helical segments are arranged relative to each other at an angle of ca. 142°. The head-to-tail cyclization increases amphipathicity of the peptide, this, however, does not affect its antimicrobial potency. Copyright © 2014 European Peptide Society and John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

The oral cavity is a primary target for opportunistic infections, particularly oral candidiasis caused by the opportunistic pathogen Candida albicans. HIV+ individuals constitute a population highly susceptible to oral candidiasis possibly due to a change in the environment of the oral cavity as the result of salivary gland dysfuntion. Histatins are a family of salivary antimicrobial peptides which under normal circumstances have a protective function on the oral mucosa. This study aimed to compare salivary histatin concentrations and oral fungal colonisation in an HIV+ and HIV− control populations. Oral samples for fungal cultures and parotid saliva were collected from all subjects. Fungal identification was determined using standard mycological procedures. In order to determine salivary histatin levels a semi-quantitative ELISA was designed using a specific polyclonal antibody and extensive statistical analysis was performed. Forty-seven percent of HIV+ and 17% of control subjects had positive fungal cultures. Mean histatin levels were 7.32 μg ml−1 for the HIV+ group and 9.17 μg ml−1 for control group ( P = 0.003). The data from this study demonstrate that the level of fungal colonisation is significantly higher in HIV+ individuals whereas histatin-5 concentrations are significantly lower, likely contributing to the enhanced predisposition of this population to oral candidiasis. [ABSTRACT FROM AUTHOR]

Abstract: The results of 520 ps molecular dynamics simulation of histatin-5, a small peptide present in human saliva and possessing antimicrobial activity, dissolved in water and in 2,2,2-trifluoroethanol, are reported. The simulations indicate that histatin-5 is destabilized in water and begins to unfold after 250 ps, while in organic solvent it maintains a regular secondary structure throughout the trajectory. Analysis of the peptide-solvent hydrogen bonds indicates that 2,2,2-trifluoroethanol is a poorer proton acceptor than water. The fluorine atom of the alcohol is almost never engaged in a hydrogen bond and the organic solvent interacts mainly with the peptide through its hydroxyl group. For some residues analysis of the solvent residence time indicated longer values for 2,2,2-trifluoroethanol than for water. The most striking difference is related to the number of times the solvent enters and leaves the first coordination shell of the peptide. This value was more than one order of magnitude higher for water than for the alcohol, suggesting that this may be the main cause of α-helix destabilization perpetrated by water. [ABSTRACT FROM AUTHOR]

Candida albicans is a common microorganism of human's microbiota and can be easily found in both respiratory and gastrointestinal tracts as well as in the genitourinary tract. Approximately 30% of people will be infected by C. albicans during their lifetime. Due to its easy adaptation, this microorganism started to present high resistance to antifungal agents which is associated with their indiscriminate use. There are several reports of adaptive mechanisms that this species can present. Some of them are intrinsic alteration in drug targets, secretion of extracellular enzymes to promote host protein degradation and efflux receptors that lead to a diminished action of common antifungal and host's innate immune response. The current review aims to bring promising alternatives for the treatment of candidiasis caused mainly by C. albicans. One of these alternatives is the use of antifungal peptides (AFPs) from the Histatin family, like histatin-5. Besides that, our focus is to show how nanotechnology can allow the application of these peptides for treatment of this microorganism. In addition, our intention is to show the importance of nanoparticles (NPs) for this purpose, which may be essential in the near future. [ABSTRACT FROM AUTHOR]

Fungicidal effects of histatin-5 against 26 oral isolates belonging to 5 non-albicans Candida species were examined. Fifty μM of histatin-5 killed more than 95% of Candida tropicalis and Candida guilliermondii isolates and more than 90% of Candida parapsilosis and Candida krusei. However, Candida glabrata was less sensitive to the peptide (mean 62.9%). Our results, taken together, demonstrated that histatin-5 possessed the fungicidal activity against Candida species other than C. glabrata. [ABSTRACT FROM AUTHOR]

Antimicrobial peptides (AMPs) have potential antifungal activities; however, their intracellular protein targets are poorly reported. Proteome microarray is an effective tool with high-throughput and rapid platform that systematically identifies the protein targets. In this study, we have used yeast proteome microarrays for systematical identification of the yeast protein targets of Lactoferricin B (Lfcin B) and Histatin-5. A total of 140 and 137 protein targets were identified from the triplicate yeast proteome microarray assays for Lfcin B and Histatin-5, respectively. The Gene Ontology (GO) enrichment analysis showed that Lfcin B targeted more enrichment categories than Histatin-5 did in all GO biological processes, molecular functions, and cellular components. This might be one of the reasons that Lfcin B has a lower minimum inhibitory concentration (MIC) than Histatin-5. Moreover, pairwise essential proteins that have lethal effects on yeast were analyzed through synthetic lethality. A total of 11 synthetic lethal pairs were identified within the protein targets of Lfcin B. However, only three synthetic lethal pairs were identified within the protein targets of Histatin-5. The higher number of synthetic lethal pairs identified within the protein targets of Lfcin B might also be the reason for Lfcin B to have lower MIC than Histatin-5. Furthermore, two synthetic lethal pairs were identified between the unique protein targets of Lfcin B and Histatin-5. Both the identified synthetic lethal pairs proteins are part of the Spt-Ada-Gcn5 acetyltransferase (SAGA) protein complex that regulates gene expression via histone modification. Identification of synthetic lethal pairs between Lfcin B and Histatin-5 and their involvement in the same protein complex indicated synergistic combination between Lfcin B and Histatin-5. This hypothesis was experimentally confirmed by growth inhibition assay. [ABSTRACT FROM AUTHOR]