Calcitermin

Calcitermin, an antimicrobial peptide from the fluid of the human airways, is a well-conserved, 15 amino acid C-terminal cleavage fragment of calgranulin C (VAIALKAAHYHTHKE), which is active under acidic pH conditions (pH 5.4). In an attempt to understand the impact of the coordination of Zn(ii) and Cu(ii) on the biological activity of calcitermin, we mutated each of the histidines with an alanine and studied the thermodynamics, binding mode and antimicrobial activity of wild type calcitermin and its H9A, H11A and H13A mutants and their Zn(ii) and Cu(ii) complexes. Both metals strongly enhance the antimicrobial activity of calcitermin-like peptides, although the link between the minimal inhibitory concentration (MIC) values and the stability, charge or structure of the complexes is not so obvious. As expected, the increase in the number of histidines makes the coordination of both metals more effective. There is no preferred Cu(ii) binding site in calcitermin: the stabilities of the Cu(ii)-H9A and Cu(ii)-H13A complexes are almost identical, while the Cu(ii)-H11A complex (in which two histidines are separated by three amino acids and only one His residue is involved in binding) is less stable. On the other hand, the higher stability of the Zn(ii)-H13A complex with respect to those formed by H9A and H11A suggests a pivotal role of His9 and His11 in Zn(ii) complexation. Impressive MIC breakpoints were obtained, similar and lower than those for commonly used antimicrobial agents that treat Candida albicans (Zn(ii) and Cu(ii) complexes of WT calcitermin and H9A, as well as H9A alone), Enterococcus faecalis (H11A, H13A and their metal complexes) and Staphylococcus aureus (H13A and its complexes).

Calcitermin is an antimicrobial peptide of 15 amino acids found in human nasal fluid characterized by antifungal and antibacterial properties. Candida albicans is the most common human fungal pathogen affecting many tissues, such as vaginal mucosa. In this study a formulation suitable for calcitermin administration on vaginal mucosa was developed for the treatment of fungal infections. To favor topical application, mucosal adhesion, and permanence, gels based on poloxamer 407 and xanthan gum were designed and compared with regard to their rheological behavior, erosion, and leakage. The selected gel was loaded with calcitermin, whose release kinetic was evaluated in vitro by Franz cells. An antifungal activity assay was conducted to assess the calcitermin anticandidal potential and the effect of its inclusion in the selected gel. The rheological study revealed the elastic and viscous moduli behavior as a function of poloxamer 407 and xanthan gum concentration. Xanthan gum presence decreased the transition temperature of the gel, while prolonging its erosion and leakage. Particularly, poloxamer 407, 18% and xanthan gum 0.4% were chosen. The calcitermin loading in the selected gel resulted in a transparent and homogeneous formulation and in a 4-fold decrease of the release rate with respect to the calcitermin solution, as evidenced by Franz cell study. The anticandidal activity tests demonstrated that calcitermin-loaded gel was more active against Candida albicans with respect to the peptide solution.

The human airways are protected from pathogenic colonization by a blanket of fluid impregnated with innate antimicrobial effector molecules. Among several previously uncharacterized components, we isolated a peptide that had activity primarily targeting Gram-negative bacteria. We named the peptide 'calcitermin' since its amino acid sequence and mass were equivalent to the 15 C-terminal residues of the S100 protein, calgranulin C. The antimicrobial activity of calcitermin was enhanced in acidic buffers (pH 5.4) and in the presence of micromolar concentrations of ZnCl(2). Analysis revealed a putative zinc-binding consensus sequence as well as an alpha-helical conformation in structure-promoting solvents.