Ovotransferrin and Its γ-Core-Containing Peptides Impair Pma1-Dependent Proton Homeostasis in Candida albicans

Ovotransferrin (OvoTf) is an iron-binding glycoprotein of avian innate immunity with antimicrobial activity, but its antifungal mechanism remains incompletely understood. We examined the candidacidal activity of OvoTf and two derived γ-core-containing peptides, kaliocin-3 (Kal-3) and kaliocin-4 (Kal-4), against Candida albicans. All three agents reduced fungal viability in a concentration-dependent manner, with minimal propidium iodide uptake, indicating a predominantly non-permeabilizing mechanism. Their activity was attenuated by extracellular Na⁺ and K⁺, whereas isoosmotic sorbitol did not confer protection. OvoTf induced partial K⁺ release, and tetraethylammonium increased cell survival. Inhibition of mitochondrial complex I or ATP synthase also reduced susceptibility, although none of the agents detectably suppressed oxygen consumption under the conditions tested. OvoTf, Kal-3, and Kal-4 caused plasma membrane depolarization, ATP accumulation, and impaired glucose-induced proton extrusion. This phenotype is consistent with disruption of Pma1-dependent proton and ion homeostasis, with mitochondrial bioenergetic activity contributing as a downstream requirement rather than constituting the primary target. Neither peptide caused detectable hemolysis; Kal-4 had minimal effects on mammalian cell metabolic viability, whereas Kal-3 reduced MCF-7 metabolic viability at higher concentrations. The activity of γ-core regions in both OvoTf lobes supports their further study as templates for mechanism-guided antifungal design.