Studies of the mechanism of human salivary histatin-5 candidacidal activity with histatin-5 variants and azole-sensitive and -resistant Candida species

Histatins are a group of small, cationic, antifungal peptides present in human saliva, A previous molecular modeling analysis suggested structural similarity between the Phe(14)-His(15) and His(18)-His(19) dipeptide sequences in histatin-5 (Hsn-5; a 24-amino-acid polypeptide) and the sequence of miconazole (one of the stole-based antifungal therapeutic agents), implying that the mechanisms of killing of Candida albicans by these two molecules may be similar, To further elaborate on this observation, we have produced two variants of Hsn-5 in which Phe(14)-His(15) or His(18)-His(19) dipeptide sequences were replaced by Ala-Ala (F14A/H15A and H18A/H19A) to eliminate the phenyl and imidazole rings of the side chains and assessed their candidacidal activities against C, albicans, In addition, we tested azole-resistant C, albicans and Candida glabrata strains for their susceptibilities to Hsn-5. Analysis of the purified recombinant proteins for their candidacidal activities indicated that both variants were significantly less effective (the molar concentrations required to kill half of the maximum number of cells [ED50,s], similar to 67 and similar to 149 mu M for F14A/H15A and H18A/H19A, respectively) than the unaltered Hsn-5 (ED50, similar to 8 mu M) at killing C. albicans, suggesting that the two dipeptide sequences are important for the candidacidal activity of Hsn-5, Assessment of the candidacidal activity of Hsn-5 with the well-characterized azole-resistant strains of C, albicans and C. glabrata, however, suggested that the mode of action of histatins against Candida is distinct from that of azole-based antifungal agents because Hsn-5 kills both azole-sensitive and azole-resistant strains equally well.