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Kinetic characterisation and inhibitor sensitivity of Candida albicans and Candida auris recombinant AOX expressed in a self-assembled proteoliposome system.

ABSTRACT: Candidemia caused by Candida spp. is a serious threat in hospital settings being a major cause of acquired infection and death and a possible contributor to Covid-19 mortality. Candidemia incidence has been rising worldwide following increases in fungicide-resistant pathogens highlighting the need for more effective antifungal agents with novel modes of action. The membrane-bound enzyme alternative oxidase (AOX) promotes fungicide resistance and is absent in humans making it a desirable therapeutic target. However, the lipophilic nature of the AOX substrate (ubiquinol-10) has hindered its kinetic characterisation in physiologically-relevant conditions. Here, we present the purification and expression of recombinant AOXs from C. albicans and C. auris in a self-assembled proteoliposome (PL) system. Kinetic parameters (K_m and V_max) with respect to ubiquinol-10 have been determined. The PL system has also been employed in dose-response assays with novel AOX inhibitors. Such information is critical for the future development of novel treatments for Candidemia.

SUBMITTER: Copsey AC

PROVIDER: S-EPMC8292455 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Candida auris is a recently described multidrug-resistant pathogenic fungus that is increasingly responsible for health care-associated outbreaks across the world. Bloodstream infections of this fungus cause death in up to 70% of cases. Aggravating this scenario, the disease-promoting mechanisms of C. auris are poorly understood. Fungi release extracellular vesicles (EVs) that carry a broad range of molecules, including proteins, lipids, carbohydrates, pigments, and RNA, many of which are virulence factors. Here, we carried out a comparative molecular characterization of C. auris and Candida albicans EVs and evaluated their capacity to modulate effector mechanisms of host immune defense. Using proteomics, lipidomics, and transcriptomics, we found that C. auris released EVs with payloads that were significantly different from those of EVs released by C. albicans. EVs released by C. auris potentiated the adhesion of this yeast to an epithelial cell monolayer, while EVs from C. albicans had no effect. C. albicans EVs primed macrophages for enhanced intracellular yeast killing, whereas C. auris EVs promoted survival of the fungal cells. Moreover, EVs from both C. auris and C. albicans induced the activation of bone marrow-derived dendritic cells. Together, our findings show distinct profiles and properties of EVs released by C. auris and by C. albicans and highlight the potential contribution of C. auris EVs to the pathogenesis of this emerging pathogen. IMPORTANCE Candida auris is a recently described multidrug-resistant pathogenic fungus that is responsible for outbreaks across the globe, particularly in the context of nosocomial infections. Its virulence factors and pathogenesis are poorly understood. Here, we tested the hypothesis that extracellular vesicles (EVs) released by C. auris are a disease-promoting factor. We describe the production of EVs by C. auris and compare their biological activities against those of the better-characterized EVs from C. albicans. C. auris EVs have immunoregulatory properties, of which some are opposite those of C. albicans EVs. We also explored the cargo and structural components of those vesicles and found that they are remarkably distinct compared to EVs from C. auris's phylogenetic relative Candida albicans.

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Kinetic characterisation and inhibitor sensitivity of Candida albicans and Candida auris recombinant AOX expressed in a self-assembled proteoliposome system.

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