Probing the role of the Lysine deacetylase CgRpd3p as a novel antifungal target through structure / function analysis

Site-directed mutagenesis of predicted yeast-specific residues revealed that alanine substitutions disrupted deacetylase activity and phenocopied the cg_rpd3Δ mutant, including impaired biofilm viability, reduced virulence in the Galleria mellonella infection model, and increased fluconazole susceptibility. These findings validate the structural model and identify fungal-specific residues that represent potential druggable sites for selective inhibition.

Virtual screening of 1,540 known KDAC inhibitors identified 13 fungal-specific compounds with preferential binding to CgRpd3p over HsHDAC2. Proof-of-principle experiments showed that selected compounds inhibited deacetylase activity in vitro, sensitized N. glabrata to fluconazole, and enhanced infection clearance in vivo, supporting a synergistic therapeutic strategy. Finally, given that Rpd3p functions within a multi-subunit complex, this study examined hypothesised complex members in N. glabrata.

Deletion of cgSDS3 phenocopied cg_rpd3Δ, including reduced virulence and fluconazole sensitivity. As CgSds3p is less conserved with human homologs, it represents a promising alternative antifungal target. Overall, this work establishes CgRpd3p and its associated complex as viable, selective antifungal targets to enhance fluconazole efficacy and expand the antifungal arsenal.<br><br><i>Thesis is embargoed until 31 July 2027.</i>