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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]

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]

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]