title: Iron binding modulates candidacidal properties of salivary histatin 5.
abstract: Salivary protein histatin 5 (Hst 5) is fungicidal toward Candida albicans, the causative agent of oropharyngeal candidiasis. However, its activity in saliva is compromised by salivary protease-mediated degradation and interaction with salivary salts. Hst 5 has also been shown to bind various metals in saliva-namely, Zn, Cu, and Ni. Surprisingly, interactions of Hst 5 with Fe have not been studied, although iron is one of the most abundant metals present in saliva. Using circular dichroism, we show that Hst 5 can bind up to 10 equivalents of iron as measured by loss of its alpha-helical secondary structure that is normally observed for it in trifluoroethylene. A significant decrease in the candidacidal ability of Hst 5 was observed upon iron binding, with increasing iron concentrations being inversely proportional to Hst 5 killing activity. Binding assays showed that the decrease in killing was likely a result of reduced binding (10-fold reduction) of Fe-Hst 5 to C. albicans cells. Protease stability analysis showed that Fe-Hst 5 was completely resistant to trypsin digestion. In contrast, zinc binding had limited effects on Hst 5 fungicidal activity or protease susceptibility. RNA sequencing results identified changes in iron uptake genes in Hst 5-treated C. albicans cells. Our findings thus suggest that consequences of Hst 5 binding iron not only affect candidacidal ability and proteolyic stability of Hst 5, but may also contribute to a novel killing mechanism involving interference with cellular iron metabolism.
authors.affiliation: Department of Oral Biology, School of Dental Medicine, University at Buffalo, Buffalo, New York, USA.
authors.forename: S
authors.initials: S
authors.lastname: Puri
authors.affiliation: Department of Oral Biology, School of Dental Medicine, University at Buffalo, Buffalo, New York, USA.
authors.forename: R
authors.initials: R
authors.lastname: Li
authors.affiliation: Department of Pharmaceutical Sciences, University at Buffalo, Buffalo, New York, USA.
authors.forename: D
authors.initials: D
authors.lastname: Ruszaj
authors.affiliation: Department of Oral Biology, School of Dental Medicine, University at Buffalo, Buffalo, New York, USA.
authors.forename: S
authors.initials: S
authors.lastname: Tati
authors.affiliation: Department of Oral Biology, School of Dental Medicine, University at Buffalo, Buffalo, New York, USA edgerto@buffalo.edu.
authors.forename: M
authors.initials: M
authors.lastname: Edgerton
mesh_terms: D000935:Antifungal Agents / Q000378:metabolism* / Q000494:pharmacology; D002176:Candida albicans / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D002473:Cell Wall / Q000187:drug effects; D002942:Circular Dichroism; D038181:FMN Reductase / Q000187:drug effects; D005800:Genes, Fungal / Q000187:drug effects / Q000235:genetics; D054808:Histatins / Q000378:metabolism* / Q000494:pharmacology; D006801:Humans; D007501:Iron / Q000378:metabolism*; D033902:Periplasmic Binding Proteins / Q000187:drug effects; D017433:Protein Structure, Secondary; D017423:Sequence Analysis, RNA; D014357:Trypsin / Q000378:metabolism; D015032:Zinc / Q000378:metabolism
chemical_list: D000935:Antifungal Agents; C068075:HTN3 protein, human; D054808:Histatins; D033902:Periplasmic Binding Proteins; C062885:sfu proteins, Serratia marcescens; D007501:Iron; D038181:FMN Reductase; C023903:ferric citrate iron reductase; D014357:Trypsin; D015032:Zinc
keywords: Candida albicans; antimicrobial peptides; proteolysis; saliva; transition elements; zinc